Methods and compounds for treating paramyxoviridae virus infections

ABSTRACT

Provided are methods for treating Paramyxoviridae virus infections by administering ribosides, riboside phosphates and prodrugs thereof, of Formula I: 
     
       
         
         
             
             
         
       
     
     wherein the 1′ position of the nucleoside sugar is substituted. The compounds, compositions, and methods provided are particularly useful for the treatment of Human parainfluenza and Human respiratory syncytial virus infections.

CROSS-REFERENCE TO RELATED APPLICATIONS

This Application is a Continuation of U.S. patent application Ser. No.16/042,085, filed on Jul. 23, 2018, which is a Divisional of U.S. patentapplication Ser. No. 14/613,719, filed on Feb. 4, 2015, now U.S. Pat.No. 10,065,958, issued on Sep. 4, 2018, which is a Continuation of U.S.patent application Ser. No. 13/189,373, filed on Jul. 22, 2011,abandoned, which claims the benefit of U.S. Provisional

Application 61/366,609 filed on Jul. 22, 2010.

FIELD OF THE INVENTION

The invention relates generally to methods and compounds for treatingParamyxoviridae virus infections, particularly methods and nucleosidesfor treating respiratory syncytial virus infections and parainfluenzavirus infections.

BACKGROUND OF THE INVENTION

Paramyxoviruses of the Paramyxoviridae family are negative-sense,single-stranded, RNA viruses that are responsible for many prevalenthuman and animal diseases. These viruses comprise at least two majorsubfamilies, Paramyxovirinae and Pneumovirinae. The subfamilyParamyxovirina includes the human parainfluenza viruses (HPIV), measlesvirus and mumps virus. Although, vaccines are available to preventmeasles and mumps infections, these infections caused 745, 00 deaths in2001 so additional treatments would be desireable for susceptiblepopulations. HPIV are the second most common cause of lower respiratorytract infection in younger children and collectively cause about 75% ofthe cases of Croup(http://www.cdc.gov/ncidod/dvrd/revb/respiratory/hpivfeat.htm). HPIVscan cause repeated infections throughout life including upperrespiratory tract illness and even serious lower respiratory tractdisease (e.g., pneumonia, bronchitis, and bronchiolitis), the latterbeing especially of concern among the elderly, and among patients withcompromised immune systems (Sable, Infect. Dis. Clin. North Am. 1995, 9,987-1003). Currently, no vaccines are available to prevent HPIVinfections. Therefore there is a need for anti-Paramyxovirinatherapeutics.

The subfamily Pneumovirinae includes Human respiratory syncytial virus(HRSV). Almost all children will have had an HRSV infection by theirsecond birthday. HRSV is the major cause of lower respiratory tractinfections in infancy and childhood with 0.5% to 2% of those infectedrequiring hospitalization. The elderly and adults with chronic heart,lung disease or those that are immunosuppressed also have a high riskfor developing severe HRSV disease (http://www.cdc.gov/rsv/index.html).No vaccine to prevent HRSV infection is currently available. Themonoclonal antibody palivizumab is available for infants at high risk,e.g., premature infants or those with either cardiac or lung disease,but the cost for general use is often prohibitive. Ribavirin has alsobeen used to treat HRSV infections but has limited efficacy. Therefore,there is a need for anti-Pneumovirinae therapeutics andanti-Paramyxoviridae therapeutics in general.

Ribosides of the nucleobases pyrrolo[1,2-f][1,2,4]triazine,imidazo[1,5-f][1,2,4]triazine, imidazo[1,2-f][1,2,4]triazine, and[1,2,4]triazolo[4,3-f][1,2,4]triazine have been disclosed inCarbohydrate Research 2001, 331(1), 77-82; Nucleosides & Nucleotides(1996), 15(1-3), 793-807; Tetrahedron Letters (1994), 35(30), 5339-42;Heterocycles (1992), 34(3), 569-74; J. Chem. Soc. Perkin Trans. 1 1985,3, 621-30; J. Chem. Soc. Perkin Trans. 1 1984, 2, 229-38; WO 2000056734;Organic Letters (2001), 3(6), 839-842; J. Chem. Soc. Perkin Trans. 11999, 20, 2929-2936; and J. Med. Chem. 1986, 29(11), 2231-5. Ribosidesof pyrrolo[1,2-f][1,2,4]triazine nucleobases with antiviral, anti-HCV,and anti-RdRp activity have been disclosed by Babu (WO2008/089105 andWO2008/141079) and Francom (WO2010/002877).

Butler, et al., WO2009132135, disclose 1′ substituted ribosides andprodrugs comprising pyrrolo[1,2-f][1,2,4]triazine nucleobases which haveanti-HCV and anti-RdRp activity. However, no methods of treatingParamyxoviridae infections with these compounds have been disclosed.

SUMMARY OF THE INVENTION

Provided are methods and compounds for the treatment of infections casedby the Paramyxoviridae virus family.

Provided, is a method for treating a Paramyxoviridae infection in amammal in need thereof comprising administering a therapeuticallyeffective amount of a compound of Formula I:

or a pharmaceutically acceptable salt or ester, thereof;

wherein:

each R¹ is H or halogen;

each R², R³ or R⁵ is independently H, OR^(a), N(R^(a))₂, N₃, CN, NO₂,S(O)_(n)R^(a), halogen, (C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl,(C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl, (C₂-C₈)substituted alkenyl,(C₂-C₈)alkynyl or (C₂-C₈)substituted alkynyl;

or any two R², R³ or R⁵ on adjacent carbon atoms when taken together are—O(CO)O— or when taken together with the ring carbon atoms to which theyare attached form a double bond;

R⁶ is OR^(a), N(R^(a))₂, N₃, CN, NO₂, S(O)_(n)R^(a), —C(═O)R¹¹,—C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), —SO₂NR¹¹R¹², halogen, (C₁-C₈)alkyl,(C₄-C₈)carbocyclylalkyl, (C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl,(C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, (C₂-C₈)substituted alkynyl,or aryl(C₁-C₈)alkyl;

each n is independently 0, 1, or 2;

each R^(a) is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl,(C₂-C₈)alkynyl, aryl(C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl, —C(═O)R¹¹,—C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), or —SO₂NR¹¹R¹²;

R⁷ is H, —C(═O)R¹¹, —C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹,—S(O)₂R¹¹, —S(O)(OR¹¹), —S(O)₂(OR¹¹), —SO₂NR¹¹R¹², or

each Y or Y¹ is, independently, O, S, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR), orN—NR₂;

W¹ and W², when taken together, are —Y³(C(R^(y))₂)₃Y³—; or one of W¹ orW² together with either R³ or R⁴ is —Y³— and the other of W¹ or W² isFormula Ia; or W¹ and W² are each, independently, a group of the FormulaIa:

wherein:

each Y² is independently a bond, O, CR₂, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR),N—NR₂, S, S—S, S(O), or S(O)₂;

each Y³ is independently O, S, or NR;

M2 is 0, 1 or 2;

each R^(x) is independently R^(y) or the formula:

wherein:

each M1a, M1c, and M1d is independently 0 or 1;

M12c is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12;

each R^(y) is independently H, F, Cl, Br, I, OH, R, —C(═Y¹)R, —C(═Y¹)OR,—C(═Y¹)N(R)₂, —N(R)₂, —⁺N(R)₃, —SR, —S(O)R, —S(O)₂R, —S(O)(OR),—S(O)₂(OR), —OC(═Y¹)R, —OC(═Y¹)OR, —OC(═Y¹)(N(R)₂), —SC(═Y¹)R,—SC(═Y¹)OR, —SC(═Y¹)(N(R)₂), —N(R)C(═Y¹)R, —N(R)C(═Y¹)OR,—N(R)C(═Y¹)N(R)₂, SO₂NR₂, —CN, —N₃, —NO₂, —OR, or W³; or when takentogether, two R^(y) on the same carbon atom form a carbocyclic ring of 3to 7 carbon atoms;

each R is independently H, (C₁-C₈) alkyl, (C₁-C₈) substituted alkyl,(C₂-C₈)alkenyl, (C₂-C₈) substituted alkenyl, (C₂-C₈) alkynyl, (C₂-C₈)substituted alkynyl, C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₂-C₂₀heterocyclyl, C₂-C₂₀ substituted heterocyclyl, arylalkyl or substitutedarylalkyl;

W³ is W⁴ or W⁵; W⁴ is R, —C(Y¹)R^(y), —C(Y¹)W⁵, —SO₂R^(y), or —SO₂W⁵;and W⁵ is a carbocycle or a heterocycle wherein W⁵ is independentlysubstituted with 0 to 3 R^(y) groups;

each R⁸ is halogen, NR¹¹R¹², N(R¹¹)OR¹¹, NR¹¹NR¹¹R¹², N₃, NO, NO₂, CHO,CN, —CH(═NR¹¹), —CH═NNHR¹¹, —CH═N(OR¹¹), —CH(OR¹¹)₂, —C(═O)NR¹¹R¹²,—C(═S)NR¹¹R¹², —C(═O)OR¹¹, (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl,(C₄-C₈)carbocyclylalkyl, optionally substituted aryl, optionallysubstituted heteroaryl, —C(═O)(C₁-C₈)alkyl, —S(O)_(n)(C₁-C₈)alkyl,aryl(C₁-C₈)alkyl, OR¹¹ or SR¹¹;

each R⁹ or R¹⁰ is independently H, halogen, NR¹¹R¹², N(R¹¹)OR¹¹,NR¹¹NR¹¹R¹², N₃, NO, NO₂, CHO, CN, —CH(═NR¹¹), —CH═NHNR¹¹, —CH═N(OR¹¹),—CH(OR¹¹)₂, —C(═O)NR¹¹R¹², —C(═S)NR¹¹R¹², —C(═O)OR¹¹, R¹¹, OR¹¹ or SR¹¹;

each R¹¹ or R¹² is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl,(C₂-C₈)alkynyl, (C₄-C₈)carbocyclylalkyl, optionally substituted aryl,optionally substituted heteroaryl, —C(═O)(C₁-C₈)alkyl,—S(O)_(n)(C₁-C₈)alkyl or aryl(C₁-C₈)alkyl; or R¹¹ and R¹² taken togetherwith a nitrogen to which they are both attached form a 3 to 7 memberedheterocyclic ring wherein any one carbon atom of said heterocyclic ringcan optionally be replaced with —O—, —S— or —NR^(a)—; and

wherein each (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl oraryl(C₁-C₈)alkyl of each R², R³, R⁵, R⁶, R¹¹ or R¹² is, independently,optionally substituted with one or more halo, hydroxy, CN, N₃, N(R^(a))₂or OR^(a); and wherein one or more of the non-terminal carbon atoms ofeach said (C₁-C₈)alkyl may be optionally replaced with —O—, —S— or—NR^(a)—.

In another embodiment, the method comprises administering atherapeutically effective amount of a racemate, enantiomer,diastereomer, tautomer, polymorph, pseudopolymorph, amorphous form,hydrate or solvate of a compound of Formula I or a pharmaceuticallyacceptable salt or ester thereof to a mammal in need thereof.

In another embodiment, the method comprises treating a Paramyxovirinainfection in a mammal in need thereof by administering a therapeuticallyeffective amount of a compound of Formula I or a pharmaceuticallyacceptable salt or ester thereof.

In another embodiment, the method comprises treating a parainfluenza,measles or mumps virus infection in a mammal in need thereof byadministering a therapeutically effective amount of a compound ofFormula I or a pharmaceutically acceptable salt or ester thereof.

In another embodiment, the method comprises treating a parainfluenzavirus infection in a mammal in need thereof by administering atherapeutically effective amount of a compound of Formula I or apharmaceutically acceptable salt or ester thereof.

In another embodiment, the method comprises treating a Pneumovirinaeinfection in a mammal in need thereof by administering a therapeuticallyeffective amount of a compound of Formula I or a pharmaceuticallyacceptable salt or ester thereof.

In another embodiment, the method comprises treating a respiratorysyncytial virus infection in a mammal in need thereof by administering atherapeutically effective amount of a compound of Formula I or apharmaceutically acceptable salt or ester thereof.

In another embodiment, the method comprises administering atherapeutically effective amount of a pharmaceutical compositioncomprising an effective amount of a Formula I compound, or apharmaceutically acceptable salt or ester thereof, in combination with apharmaceutically acceptable diluent or carrier.

In another embodiment, the method comprises administering atherapeutically effective amount of a pharmaceutical compositioncomprising an effective amount of a Formula I compound, or apharmaceutically acceptable salt or ester thereof, in combination withat least one additional therapeutic agent.

In another embodiment, the method comprises administering atherapeutically effective amount of a combination pharmaceutical agentcomprising:

a) a first pharmaceutical composition comprising a compound of FormulaI; or a pharmaceutically acceptable salt, solvate, or ester thereof; and

b) a second pharmaceutical composition comprising at least oneadditional therapeutic agent active against infectious Paramyxoviridaeviruses.

In another embodiment, the present application provides for a method ofinhibiting a Paramyxoviridae RNA-dependent RNA polymerase, comprisingcontacting a cell infected with a Paramyxoviridae virus with aneffective amount of a compound of Formula I; or a pharmaceuticallyacceptable salts, solvate, and/or ester thereof.

In another embodiment, provided is the use of a compound of Formula I ora pharmaceutically acceptable salt, solvate, and/or ester thereof totreat a viral infection caused by a Paramyxoviridae virus.

In another aspect, the invention also provides processes and novelintermediates disclosed herein which are useful for preparing Formula Icompounds of the invention.

In other aspects, novel methods for synthesis, analysis, separation,isolation, purification, characterization, and testing of the compoundsof this invention are provided.

DETAILED DESCRIPTION OF EXEMPLARY EMBODIMENTS

Reference will now be made in detail to certain embodiments of theinvention, examples of which are illustrated in the accompanyingdescription, structures and formulas. While the invention will bedescribed in conjunction with the enumerated embodiments, it will beunderstood that they are not intended to limit the invention to thoseembodiments. On the contrary, the invention is intended to cover allalternatives, modifications, and equivalents, which may be includedwithin the scope of the present invention.

In another embodiment, provided is a method of treating aParamyxoviridae infection in a mammal in need thereof comprisingadministering a therapeutically effective amount of a compound ofFormula I represented by Formula II:

or a pharmaceutically acceptable salt or ester, thereof;

wherein:

each R¹ is H or halogen;

each R² is OR^(a) or halogen;

each R³ or R⁵ is independently H, OR^(a), N(R^(a))₂, N₃, CN, NO₂,S(O)_(n)R^(a), halogen, (C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl,(C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl, (C₂-C₈)substituted alkenyl,(C₂-C₈)alkynyl or (C₂-C₈)substituted alkynyl;

or any two R², R³ or R⁵ on adjacent carbon atoms when taken together are—O(CO)O— or when taken together with the ring carbon atoms to which theyare attached form a double bond;

R⁶ is OR^(a), N(R^(a))₂, N₃, CN, S(O)_(n)R^(a), —C(═O)R¹¹, —C(═O)OR¹¹,—C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), —SO₂NR¹¹R¹², halogen, (C₁-C₈)alkyl,(C₄-C₈)carbocyclylalkyl, (C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl,(C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, or (C₂-C₈)substitutedalkynyl;

each n is independently 0, 1, or 2;

each R^(a) is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl,(C₂-C₈)alkynyl, aryl(C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl, —C(═O)R¹¹,—C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), or —SO₂NR¹¹R¹²;

R⁷ is H, —C(═O)R¹¹, —C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹,—S(O)₂R¹¹, —S(O)(OR¹¹), —S(O)₂(OR¹¹), —SO₂NR¹¹R¹², or

each Y or Y¹ is, independently, O, S, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR), orN—NR₂;

W¹ and W², when taken together, are —Y³(C(R^(y))₂)₃Y³—; or one of W¹ orW² together with either R³ or R⁴ is —Y³— and the other of W¹ or W² isFormula Ia; or W¹ and W² are each, independently, a group of the FormulaIa:

wherein:

each Y² is independently a bond, O, CR₂, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR),N—NR₂, S, S—S, S(O), or S(O)₂;

each Y³ is independently O, S, or NR;

M2 is 0, 1 or 2;

each R^(x) is independently R^(y) or the formula:

wherein:

each M1a, M1c, and M1d is independently 0 or 1;

M12c is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12;

each R^(y) is independently H, F, Cl, Br, I, OH, R, —C(═Y¹)R, —C(═Y¹)OR,—C(═Y¹)N(R)₂, —N(R)₂, —⁺N(R)₃, —SR, —S(O)R, —S(O)₂R, —S(O)(OR),—S(O)₂(OR), —OC(═Y¹)R, —OC(═Y¹)OR, —OC(═Y¹)(N(R)₂), —SC(═Y¹)R,—SC(═Y¹)OR, —SC(═Y¹)(N(R)₂), —N(R)C(═Y¹)R, —N(R)C(═Y¹)OR,—N(R)C(═Y¹)N(R)₂, —SO₂NR₂, —CN, —N₃, —NO₂, —OR, or W³; or when takentogether, two R^(y) on the same carbon atom form a carbocyclic ring of 3to 7 carbon atoms;

each R is independently H, (C₁-C₈) alkyl, (C₁-C₈) substituted alkyl,(C₂-C₈)alkenyl, (C₂-C₈) substituted alkenyl, (C₂-C₈) alkynyl, (C₂-C₈)substituted alkynyl, C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₂-C₂₀heterocyclyl, C₂-C₂₀ substituted heterocyclyl, arylalkyl or substitutedarylalkyl;

W³ is W⁴ or W⁵; W⁴ is R, —C(Y¹)R^(y), —C(Y¹)W⁵, —SO₂R^(y), or —SO₂W⁵;and W⁵ is a carbocycle or a heterocycle wherein W⁵ is independentlysubstituted with 0 to 3 R^(y) groups;

each R⁸ is halogen, NR¹¹R¹², N(R¹¹)OR¹¹, NR¹¹NR¹¹R¹² N₃, NO, NO₂, CHO,CN, —CH(═NR¹¹), —CH═NNHR¹¹, —CH═N(OR¹¹), —CH(OR¹¹)₂, —C(═O)NR¹¹R¹²,—C(═S)NR¹¹R¹², —C(═O)OR¹¹, (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl,(C₄-C₈)carbocyclylalkyl, optionally substituted aryl, optionallysubstituted heteroaryl, —C(═O)(C₁-C₈)alkyl, —S(O)_(n)(C₁-C₈)alkyl,aryl(C₁-C₈)alkyl, OR¹¹ or SR¹¹;

each R⁹ is independently H, halogen, NR¹¹R¹², N(R¹¹)OR¹¹, NR¹¹NR¹¹R¹²,N₃, NO, NO₂, CHO, CN, —CH(═NR¹¹), —CH═NHNR¹¹, —CH═N(OR¹¹), —CH(OR¹¹)₂,—C(═O)NR¹¹R¹², —C(═S)NR¹¹R¹², —C(═O)OR¹¹, R¹¹, OR¹¹ or SR¹¹;

each R¹¹ or R¹² is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl,(C₂-C₈)alkynyl, (C₄-C₈)carbocyclylalkyl, optionally substituted aryl,optionally substituted heteroaryl, —C(═O)(C₁-C₈)alkyl,—S(O)_(n)(C₁-C₈)alkyl or aryl(C₁-C₈)alkyl; or R¹¹ and R¹² taken togetherwith a nitrogen to which they are both attached form a 3 to 7 memberedheterocyclic ring wherein any one carbon atom of said heterocyclic ringcan optionally be replaced with —O—, —S— or —NR^(a)—; and

wherein each (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl oraryl(C₁-C₈)alkyl of each R³, R⁵, R⁶, R¹¹ or R¹² is, independently,optionally substituted with one or more halo, hydroxy, CN, N₃, N(R^(a))₂or OR^(a); and wherein one or more of the non-terminal carbon atoms ofeach said (C₁-C₈)alkyl may be optionally replaced with —O—, —S— or—NR^(a)—.

In one embodiment of the method of treating a Paramyxoviridae infectionby administering a compound of Formula II, R¹ of Formula II is H. Inanother aspect of this embodiment R⁶ of Formula II is N₃, CN, halogen,(C₁-C₈)alkyl, (C₁-C₈) substituted alkyl, (C₂-C₈)alkenyl,(C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, or (C₂-C₈)substitutedalkynyl. In another aspect of this embodiment, R⁶ of Formula II is CN,methyl, ethenyl, or ethynyl. In another aspect of this embodiment, R⁶ ofFormula II is CN. In another aspect of this embodiment, R⁶ of Formula IIis methyl. In another aspect of this embodiment, R⁵ of Formula II is H.In another aspect of this embodiment, R² of Formula II is OR^(a). Inanother aspect of this embodiment, R² of Formula II is OH. In anotheraspect of this embodiment, R² of Formula II is F. In another aspect ofthis embodiment, R³ of Formula II is OR^(a). In another aspect of thisembodiment, R³ of Formula II is OH, —OC(═O)R¹¹, or —OC(═O)OR¹¹. Inanother aspect of this embodiment, R³ of Formula II is OH. In anotheraspect of this embodiment, R⁸ of Formula II is NR¹¹R¹². In anotheraspect of this embodiment, R⁸ of Formula II is NH₂. In another aspect ofthis embodiment, R⁸ of Formula II is OR¹¹. In another aspect of thisembodiment, R⁸ of Formula II is OH. In another aspect of thisembodiment, R⁹ of Formula II is H. In another aspect of this embodiment,R⁹ of Formula II is NR¹¹R¹². In another aspect of this embodiment, R⁹ ofFormula II is NH₂. In another aspect of this embodiment, R⁷ of FormulaII is H, —C(═O)R¹¹, —C(═O)OR¹¹ or

In another aspect of this embodiment, R⁷ of Formula II is H. In anotheraspect of this embodiment, R⁷ of Formula II is

In another embodiment of the method of treating a Paramyxoviridaeinfection comprising administering a compound of Formula II, theParamyxoviridae infection is caused by a Paramyxovirina virus. Inanother aspect of this embodiment, the Paramyxovirina virus is aparainfluenza, measles or mumps virus. In another aspect of thisembodiment, the Paramyxovirina virus is a Respirovirus virus. In anotheraspect of this embodiment, the Paramyxovirina virus is a type 1 or 3Human parainfluenza virus.

In another embodiment of the method of treating a Paramyxoviridaeinfection comprising administering a compound of Formula II, theParamyxoviridae infection is caused by a Pneumovirinae virus. In anotheraspect of this embodiment, the Pneumovirinae virus is a respiratorysyncytial virus. In another aspect of this embodiment, the Pneumovirinaevirus is a Human respiratory syncytial virus.

In another embodiment, provided is a method of treating aParamyxoviridae infection in a mammal in need thereof comprisingadministering a therapeutically effective amount of a compound ofFormula I represented by Formula III:

or a pharmaceutically acceptable salt or ester, thereof;

wherein:

each R² is OR^(a) or F;

each R³ is OR^(a);

R⁶ is OR^(a), N(R^(a))₂, N₃, CN, S(O)_(n)R^(a), —C(═O)R¹¹, —C(═O)OR¹¹,—C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), —SO₂NR¹¹R¹², halogen, (C₁-C₈)alkyl,(C₄-C₈)carbocyclylalkyl, (C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl,(C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, or (C₂-C₈)substitutedalkynyl;

each n is independently 0, 1, or 2;

each R^(a) is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl,(C₂-C₈)alkynyl, aryl(C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl, —C(═O)R¹¹,—C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), or —SO₂NR¹¹R¹²;

R⁷ is H, —C(═O)R¹¹, —C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹,—S(O)₂R¹¹, —S(O)(OR¹¹), —S(O)₂(OR¹¹), SO₂NR¹¹R¹², or

each Y or Y¹ is, independently, O, S, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR), orN—NR₂;

W¹ and W², when taken together, are —Y³(C(R^(y))₂)₃Y³—; or one of W¹ orW² together with either R³ or R⁴ is —Y³— and the other of W¹ or W² isFormula Ia; or W¹ and W² are each, independently, a group of the FormulaIa:

wherein:

each Y² is independently a bond, O, CR₂, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR),N—NR₂, S, S—S, S(O), or S(O)₂;

each Y³ is independently O, S, or NR;

M2 is 0, 1 or 2;

each R^(x) is independently R^(y) or the formula:

wherein:

each M1a, M1c, and M1d is independently 0 or 1;

M12c is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12;

each R^(y) is independently H, F, Cl, Br, I, OH, R, —C(═Y¹)R, —C(═Y¹)OR,—C(═Y¹)N(R)₂, —N(R)₂, —⁺N(R)₃, —SR, —S(O)R, —S(O)₂R, —S(O)(OR),—S(O)₂(OR), —OC(═Y¹)R, —OC(═Y¹)OR, —OC(═Y¹)(N(R)₂), —SC(═Y¹)R,—SC(═Y¹)OR, —SC(═Y¹)(N(R)₂), —N(R)C(═Y¹)R, —N(R)C(═Y¹)OR,—N(R)C(═Y¹)N(R)₂, —SO₂NR₂, —CN, —N₃, —NO₂, —OR, or W³; or when takentogether, two R^(y) on the same carbon atom form a carbocyclic ring of 3to 7 carbon atoms;

each R is independently H, (C₁-C₈) alkyl, (C₁-C₈) substituted alkyl,(C₂-C₈)alkenyl, (C₂-C₈) substituted alkenyl, (C₂-C₈) alkynyl, (C₂-C₈)substituted alkynyl, C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₂-C₂₀heterocyclyl, C₂-C₂₀ substituted heterocyclyl, arylalkyl or substitutedarylalkyl;

W³ is W⁴ or W⁵; W⁴ is R, —C(Y¹)R^(y), —C(Y¹)W⁵, —SO₂R^(y), or —SO₂W⁵;and W⁵ is a carbocycle or a heterocycle wherein W⁵ is independentlysubstituted with 0 to 3 R^(y) groups;

each R⁸ is halogen, NR¹¹R¹², N(R¹¹)OR¹¹, NR¹¹NR¹¹R¹², N₃, NO, NO₂, CHO,CN, —CH(═NR¹¹), —CH═NNHR¹¹, —CH═N(OR¹¹), —CH(OR¹¹)₂, —C(═O)NR¹¹R¹²,—C(═S)NR¹¹R¹², —C(═O)OR¹¹, (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl,(C₄-C₈)carbocyclylalkyl, optionally substituted aryl, optionallysubstituted heteroaryl, —C(═O)(C₁-C₈)alkyl, —S(O)_(n)(C₁-C₈)alkyl,aryl(C₁-C₈)alkyl, OR¹¹ or SR¹¹;

each R⁹ is independently H, halogen, NR¹¹R¹², N(R¹¹)OR¹¹, NR¹¹NR¹¹R¹²,N₃, NO, NO₂, CHO, CN, —CH(═NR¹¹), —CH═NHNR¹¹, —CH═N(OR¹¹), —CH(OR¹¹)₂,—C(═O)NR¹¹R¹², —C(═S)NR¹¹R¹², —C(═O)OR¹¹, R¹¹, OR¹¹ or SR¹¹; and

each R¹¹ or R¹² is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl,(C₂-C₈)alkynyl, (C₄-C₈)carbocyclylalkyl, optionally substituted aryl,optionally substituted heteroaryl, —C(═O)(C₁-C₈)alkyl,—S(O)_(n)(C₁-C₈)alkyl or aryl(C₁-C₈)alkyl; or R¹¹ and R¹² taken togetherwith a nitrogen to which they are both attached form a 3 to 7 memberedheterocyclic ring wherein any one carbon atom of said heterocyclic ringcan optionally be replaced with —O—, —S— or —NR^(a)—; and

wherein each (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl oraryl(C₁-C₈)alkyl of each R⁶, R¹¹ or R¹² is, independently, optionallysubstituted with one or more halo, hydroxy, CN, N₃, N(R^(a))₂ or OR^(a);and wherein one or more of the non-terminal carbon atoms of each said(C₁-C₈)alkyl may be optionally replaced with —O—, —S— or —NR^(a)—.

In one embodiment of the method of treating a Paramyxoviridae infectioncomprising administering a compound of Formula III, R⁶ of Formula III isN₃, CN, halogen, (C₁-C₈)alkyl, (C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl,(C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, or (C₂-C₈)substitutedalkynyl. In another aspect of this embodiment, R⁶ of Formula III is CN,methyl, ethenyl, or ethynyl. In another aspect of this embodiment, R⁶ ofFormula III is CN. In another aspect of this embodiment, R⁶ of FormulaIII is methyl. In another aspect of this embodiment, R² of Formula IIIis OR^(a). In another aspect of this embodiment, R² of Formula III isOH. In another aspect of this embodiment, R² of Formula III is F. Inanother aspect of this embodiment, R³ of Formula III is OH, —OC(═O)R¹¹,or —OC(═O)OR¹¹. In another aspect of this embodiment, R³ of Formula IIIis OH. In another aspect of this embodiment, R⁸ of Formula III isNR¹¹R¹². In another aspect of this embodiment, R⁸ of Formula III is NH₂.In another aspect of this embodiment, R⁸ of Formula III is OR¹¹. Inanother aspect of this embodiment, R⁸ of Formula III is OH. In anotheraspect of this embodiment, R⁹ of Formula III is H. In another aspect ofthis embodiment, R⁹ of Formula III is NR¹¹R¹². In another aspect of thisembodiment, R⁹ of Formula III is NH₂. In another aspect of thisembodiment, R⁷ of Formula III is H, —C(═O)R¹¹, —C(═O)OR¹¹ or

In another aspect of this embodiment, R⁷ of Formula III is H. In anotheraspect of this embodiment, R⁷ of Formula III is

In another embodiment of the method of treating a Paramyxoviridaeinfection comprising administering a compound of Formula III, R⁶ ofFormula III is N₃, CN, halogen, (C₁-C₈)alkyl, (C₁-C₈)substituted alkyl,(C₂-C₈)alkenyl, (C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, or(C₂-C₈)substituted alkynyl and R⁸ is NH₂. In another aspect of thisembodiment, R⁶ of Formula III is CN, methyl, ethenyl, or ethynyl. Inanother aspect of this embodiment, R⁶ of Formula III is CN. In anotheraspect of this embodiment, R⁶ of Formula III is methyl. In anotheraspect of this embodiment, R² of Formula III is OR^(a). In anotheraspect of this embodiment, R² of Formula III is OH, —OC(═O)R¹¹, or—OC(═O)OR¹¹. In another aspect of this embodiment, R² of Formula III isOH. In another aspect of this embodiment, R² of Formula III is F. Inanother aspect of this embodiment, R³ of Formula III is OH, —OC(═O)R¹¹,or —OC(═O)OR¹¹. In another aspect of this embodiment, R³ of Formula IIIis OH. In another aspect of this embodiment, R⁹ of Formula III is H. Inanother aspect of this embodiment, R⁹ of Formula III is NR¹¹R¹². Inanother aspect of this embodiment, R⁹ of Formula III is NH₂. In anotheraspect of this embodiment, R⁷ of Formula III is H, —C(═O)R¹¹, —C(═O)OR¹¹or

In another aspect of this embodiment, R⁷ of Formula III is H. In anotheraspect of this embodiment, R⁷ of Formula III is

In another embodiment of the method of treating a Paramyxoviridaeinfection comprising administering a compound of Formula III, R⁶ ofFormula III is CN, methyl, ethenyl, or ethynyl, R⁸ is NH₂, and R⁹ is H.In another aspect of this embodiment, R⁶ of Formula III is CN. Inanother aspect of this embodiment, R⁶ of Formula III is methyl. Inanother aspect of this embodiment, R² of Formula III is OR^(a). Inanother aspect of this embodiment, R² of Formula III is OH, —OC(═O)R¹¹,or —OC(═O)OR¹¹. In another aspect of this embodiment, R² of Formula IIIis OH. In another aspect of this embodiment, R² of Formula III is F. Inanother aspect of this embodiment, R³ of Formula III is OH, —OC(═O)R¹¹,or —OC(═O)OR¹¹. In another aspect of this embodiment, R³ of Formula IIIis OH. In another aspect of this embodiment, R⁷ of Formula III is H,—C(═O)R¹¹, —C(═O)OR¹¹ or

In another aspect of this embodiment, R⁷ of Formula III is H. In anotheraspect of this embodiment, R⁷ of Formula III is

In another embodiment of the method of treating a Paramyxoviridaeinfection comprising administering a compound of Formula III, theParamyxoviridae infection is caused by a Paramyxovirina virus. Inanother aspect of this embodiment, the Paramyxovirina virus is aparainfluenza, measles or mumps virus. In another aspect of thisembodiment, the Paramyxovirina virus is a Respirovirus virus. In anotheraspect of this embodiment, the Paramyxovirina virus is a type 1 or 3Human parainfluenza virus.

In another embodiment of the method of treating a Paramyxoviridaeinfection comprising administering a compound of Formula III, theParamyxoviridae infection is caused by a Pneumovirinae virus. In anotheraspect of this embodiment, the Pneumovirinae virus is a respiratorysyncytial virus. In another aspect of this embodiment, the Pneumovirinaevirus is a Human respiratory syncytial virus.

In one embodiment, provided is a compound of Formula IV:

or a pharmaceutically acceptable salt or ester, thereof;

wherein:

each R¹ is H or halogen;

each R³ or R⁵ is independently H, OR^(a), N(R^(a))₂, N₃, CN, NO₂,S(O)_(n)R^(a), halogen, (C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl,(C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl, (C₂-C₈)substituted alkenyl,(C₂-C₈)alkynyl or (C₂-C₈)substituted alkynyl;

R⁶ is OR^(a), N(R^(a))₂, N₃, CN, S(O)_(n)R^(a), —C(═O)R¹¹, —C(═O)OR¹¹,—C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), —SO₂NR¹¹R¹², halogen, (C₁-C₈)alkyl,(C₄-C₈)carbocyclylalkyl, (C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl,(C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, or (C₂-C₈)substitutedalkynyl;

each n is independently 0, 1, or 2;

each R^(a) is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl,(C₂-C₈)alkynyl, aryl(C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl, —C(═O)R¹¹,—C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), or —SO₂NR¹¹R¹²;

R⁷ is H, —C(═O)R¹¹, —C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹,—S(O)₂R¹¹, —S(O)(OR¹¹), —S(O)₂(OR¹¹), —SO₂NR¹¹R¹², or

each Y or Y¹ is, independently, O, S, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR), orN—NR₂;

W¹ and W², when taken together, are —Y³(C(R^(y))₂)₃Y³—; or one of W¹ orW² together with either R³ or R⁴ is —Y³— and the other of W¹ or W² isFormula Ia; or W¹ and W² are each, independently, a group of the FormulaIa:

wherein:

each Y² is independently a bond, O, CR₂, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR),N—NR₂, S, S—S, S(O), or S(O)₂;

each Y³ is independently O, S, or NR;

M2 is 0, 1 or 2;

each R^(x) is independently R^(y) or the formula:

wherein:

each M1a, M1c, and M1d is independently 0 or 1;

M12c is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12;

each R^(y) is independently H, F, Cl, Br, I, OH, R, —C(═Y¹)R, —C(═Y¹)OR,—C(═Y¹)N(R)₂, —N(R)₂, —⁺N(R)₃, —SR, —S(O)R, —S(O)₂R, —S(O)(OR),—S(O)₂(OR), —OC(═Y¹)R, —OC(═Y¹)OR, —OC(═Y¹)(N(R)₂), —SC(═Y¹)R,—SC(═Y¹)OR, —SC(═Y¹)(N(R)₂), —N(R)C(═Y¹)R, —N(R)C(═Y¹)OR,—N(R)C(═Y¹)N(R)₂, —SO₂NR₂, —CN, —N₃, —NO₂, —OR, or W³; or when takentogether, two R^(y) on the same carbon atom form a carbocyclic ring of 3to 7 carbon atoms;

each R is independently H, (C₁-C₈) alkyl, (C₁-C₈) substituted alkyl,(C₂-C₈)alkenyl, (C₂-C₈) substituted alkenyl, (C₂-C₈) alkynyl, (C₂-C₈)substituted alkynyl, C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₂-C₂₀heterocyclyl, C₂-C₂₀ substituted heterocyclyl, arylalkyl or substitutedarylalkyl;

W³ is W⁴ or W⁵; W⁴ is R, —C(Y¹)R^(y), —C(Y¹)W⁵, —SO₂R^(y), or —SO₂W⁵;and W⁵ is a carbocycle or a heterocycle wherein W⁵ is independentlysubstituted with 0 to 3 R^(y) groups;

each R⁸ is halogen, NR¹¹R¹², N(R¹¹)OR¹¹, NR¹¹NR¹¹R¹², N₃, NO, NO₂, CHO,CN, —CH(═NR¹¹), —CH═NNHR¹¹, —CH═N(OR¹¹), —CH(OR¹¹)₂, —C(═O)NR¹¹R¹²,—C(═S)NR¹¹R¹², —C(═O)OR¹¹, (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl,(C₄-C₈)carbocyclylalkyl, optionally substituted aryl, optionallysubstituted heteroaryl, —C(═O)(C₁-C₈)alkyl, —S(O)_(n)(C₁-C₈)alkyl,aryl(C₁-C₈)alkyl, OR¹¹ or SR¹¹;

each R⁹ is independently H, halogen, NR¹¹R¹², N(R¹¹)OR¹¹, NR¹¹NR¹¹R¹²,N₃, NO, NO₂, CHO, CN, —CH(═NR¹¹), —CH═NHNR¹¹, —CH═N(OR¹¹), —CH(OR¹¹)₂,—C(═O)NR¹¹R¹², —C(═S)NR¹¹R¹², —C(═O)OR¹¹, R¹¹, OR¹¹ or SR¹¹;

each R¹¹ or R¹² is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl,(C₂-C₈)alkynyl, (C₄-C₈)carbocyclylalkyl, optionally substituted aryl,optionally substituted heteroaryl, —C(═O)(C₁-C₈)alkyl,—S(O)_(n)(C₁-C₈)alkyl or aryl(C₁-C₈)alkyl; or R¹¹ and R¹² taken togetherwith a nitrogen to which they are both attached form a 3 to 7 memberedheterocyclic ring wherein any one carbon atom of said heterocyclic ringcan optionally be replaced with —O—, —S— or —NR^(a)—; and

wherein each (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl oraryl(C₁-C₈)alkyl of each R³, R⁵, R⁶, R¹¹ or R¹² is, independently,optionally substituted with one or more halo, hydroxy, CN, N₃, N(R^(a))₂or OR^(a); and wherein one or more of the non-terminal carbon atoms ofeach said (C₁-C₈)alkyl may be optionally replaced with —O—, —S— or—NR^(a)—.

In one embodiment of the compound of Formula IV, R⁶ is N₃, CN, halogen,(C₁-C₈)alkyl, (C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl,(C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, or (C₂-C₈)substitutedalkynyl. In another aspect of this embodiment, R⁶ is CN, methyl,ethenyl, or ethynyl. In another aspect of this embodiment, R⁶ is CN. Inanother aspect of this embodiment, R⁶ is methyl. In another aspect ofthis embodiment, R¹ is H. In another aspect of this embodiment, R³ isOH, —OC(═O)R¹¹, or —OC(═O)OR¹¹. In another aspect of this embodiment, R³is OH. In another aspect of this embodiment, R⁸ is NR¹¹R¹². In anotheraspect of this embodiment, R⁸ is NH₂. In another aspect of thisembodiment, R⁸ is OR¹¹. In another aspect of this embodiment, R⁸ is OH.In another aspect of this embodiment, R⁹ is H. In another aspect of thisembodiment, R⁹ is NR¹¹R¹². In another aspect of this embodiment, R⁹ isNH₂. In another aspect of this embodiment, R⁷ is H, —C(═O)R¹¹,—C(═O)OR¹¹ or

In another aspect of this embodiment, R⁷ is H. In another aspect of thisembodiment, R⁷ is

In another embodiment of a compound of Formula IV, R⁶ is N₃, CN,halogen, (C₁-C₈)alkyl, (C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl,(C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, or (C₂-C₈)substitutedalkynyl and R⁸ is NH₂. In another aspect of this embodiment, R⁶ is CN,methyl, ethenyl, or ethynyl. In another aspect of this embodiment, R⁶ isCN. In another aspect of this embodiment, R⁶ is methyl. In anotheraspect of this embodiment, R¹ is H. In another aspect of thisembodiment, R³ is OH, —OC(═O)R¹¹, or —OC(═O)OR¹¹. In another aspect ofthis embodiment, R³ is OH. In another aspect of this embodiment, R⁹ isH. In another aspect of this embodiment, R⁹ is NR¹¹R¹². In anotheraspect of this embodiment, R⁹ is NH₂. In another aspect of thisembodiment, R⁷ is H, —C(═O)R¹¹, —C(═O)OR¹¹ or

In another aspect of this embodiment, R⁷ is H. In another aspect of thisembodiment, R⁷ is

In another embodiment of the compound of Formula IV, R⁶ is CN, methyl,ethenyl, or ethynyl, R⁸ is NH₂, and R⁹ is H. In another aspect of thisembodiment, R¹ is H. In another aspect of this embodiment, R⁶ is CN. Inanother aspect of this embodiment, R⁶ is methyl. In another aspect ofthis embodiment, R³ is OH, —OC(═O)R¹¹, or —OC(═O)OR¹¹. In another aspectof this embodiment, R³ is OH. In another aspect of this embodiment, R⁷is H, —C(═O)R¹¹, —C(═O)OR¹¹ or

In another aspect of this embodiment, R⁷ is H. In another aspect of thisembodiment, R⁷ is

In another embodiment, provided is a method of treating aParamyxoviridae infection in a mammal in need thereof comprisingadministering a therapeutically effective amount of a compound ofFormulas I-IV, wherein R¹¹ or R¹² is independently H, (C₁-C₈)alkyl,(C₂-C₈)alkenyl, (C₂-C₈)alkynyl, (C₄-C₈)carbocyclylalkyl, optionallysubstituted aryl, optionally substituted heteroaryl, —C(═O)(C₁-C₈)alkyl,—S(O)_(n)(C₁-C₈)alkyl or aryl(C₁-C₈)alkyl. In another embodiment, R¹¹and R¹² taken together with a nitrogen to which they are both attached,form a 3 to 7 membered heterocyclic ring wherein any one carbon atom ofsaid heterocyclic ring can optionally be replaced with —O—, —S— or—NR^(a)—. Therefore, by way of example and not limitation, the moietyNR¹¹R¹² can be represented by the heterocycles:

and the like.

In another embodiment, provided is a method of treating aParamyxoviridae infection in a mammal in need thereof comprisingadministering a therapeutically effective amount of a compound ofFormula I-IV, wherein each R³, R⁴, R⁵, R⁶, R¹¹ or R¹² is, independently,(C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl or aryl(C₁-C₈)alkyl,wherein said (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl oraryl(C₁-C₈)alkyl are, independently, optionally substituted with one ormore halo, hydroxy, CN, N₃, N(R^(a))₂ or OR^(a). Therefore, by way ofexample and not limitation, R³, R⁴, R⁵, R⁶, R¹¹ or R¹² could representmoieties such as —CH(NH₂)CH₃, —CH(OH)CH2CH3, —CH(NH₂)CH(CH₃)₂, —CH₂CF₃,—(CH₂)₂CH(N₃)CH₃, —(CH₂)₆NH₂ and the like.

In another embodiment, provided is a method of treating aParamyxoviridae infection in a mammal in need thereof comprisingadministering a therapeutically effective amount of a compound ofFormula I-IV, wherein R³, R⁴, R⁵, R⁶, R¹¹ or R¹² is (C₁-C₈)alkyl whereinone or more of the non-terminal carbon atoms of each said (C₁-C₈)alkylmay be optionally replaced with —O—, —S— or —NR^(a)—. Therefore, by wayof example and not limitation, R³, R⁴, R⁵, R⁶, R¹¹ or R¹² couldrepresent moieties such as —CH₂OCH₃, —CH₂OCH₂CH₃, —CH₂OCH(CH₃)₂,—CH₂SCH₃, —(CH₂)₆OCH₃, —(CH₂)₆N(CH₃)₂ and the like.

In another embodiment, provided is a method of treating aParamyxoviridae infection in a sample comprising administering aneffective amount of a compound of Formula I selected from the groupconsisting of:

or a pharmaceutically acceptable salt or ester thereof.

In another embodiment, provided is a compound of Formula IV that is

or a pharmaceutically acceptable salt or ester thereof.

In another embodiment, provided is a compound of Formula I that is

or a pharmaceutically acceptable salt or ester thereof.

Definitions

Unless stated otherwise, the following terms and phrases as used hereinare intended to have the following meanings:

When trade names are used herein, applicants intend to independentlyinclude the tradename product and the active pharmaceuticalingredient(s) of the tradename product.

As used herein, “a compound of the invention” or “a compound of FormulaI” means a compound of Formula I or a pharmaceutically acceptable salt,thereof. Similarly, with respect to isolatable intermediates, the phrase“a compound of Formula (number)” means a compound of that formula andpharmaceutically acceptable salts, thereof.

“Alkyl” is hydrocarbon containing normal, secondary, tertiary or cycliccarbon atoms. For example, an alkyl group can have 1 to 20 carbon atoms(i.e, C₁-C₂₀ alkyl), 1 to 8 carbon atoms (i.e., C₁-C₈ alkyl), or 1 to 6carbon atoms (i.e., C₁-C₆ alkyl). Examples of suitable alkyl groupsinclude, but are not limited to, methyl (Me, —CH₃), ethyl (Et, —CH₂CH₃),1-propyl (n-Pr, n-propyl, —CH₂CH₂CH₃), 2-propyl (i-Pr, i-propyl,—CH(CH₃)₂), 1-butyl (n-Bu, n-butyl, —CH₂CH₂CH₂CH₃), 2-methyl-1-propyl(i-Bu, i-butyl, —CH₂CH(CH₃)₂), 2-butyl (s-Bu, s-butyl, —CH(CH₃)CH₂CH₃),2-methyl-2-propyl (t-Bu, t-butyl, —C(CH₃)₃), 1-pentyl (n-pentyl,—CH₂CH₂CH₂CH₂CH₃), 2-pentyl (—CH(CH₃)CH₂CH₂CH₃), 3-pentyl(—CH(CH₂CH₃)₂), 2-methyl-2-butyl (—C(CH₃)₂CH₂CH₃), 3-methyl-2-butyl(—CH(CH₃)CH(CH₃)₂), 3-methyl-1-butyl (—CH₂CH₂CH(CH₃)₂), 2-methyl-1-butyl(—CH₂CH(CH₃)CH₂CH₃), 1-hexyl (—CH₂CH₂CH₂CH₂CH₂CH₃), 2-hexyl(—CH(CH₃)CH₂CH₂CH₂CH₃), 3-hexyl (—CH(CH₂CH₃)(CH₂CH₂CH₃)),2-methyl-2-pentyl (—C(CH₃)₂CH₂CH₂CH₃), 3-methyl-2-pentyl(—CH(CH₃)CH(CH₃)CH₂CH₃), 4-methyl-2-pentyl (—CH(CH₃)CH₂CH(CH₃)₂),3-methyl-3-pentyl (—C(CH₃)(CH₂CH₃)₂), 2-methyl-3-pentyl(—CH(CH₂CH₃)CH(CH₃)₂), 2,3-dimethyl-2-butyl (—C(CH₃)₂CH(CH₃)₂),3,3-dimethyl-2-butyl (—CH(CH₃)C(CH₃)₃, and octyl (—(CH₂)₇CH₃).

“Alkoxy” means a group having the formula —O-alkyl, in which an alkylgroup, as defined above, is attached to the parent molecule via anoxygen atom. The alkyl portion of an alkoxy group can have 1 to 20carbon atoms (i.e., C₁-C₂₀ alkoxy), 1 to 12 carbon atoms(i.e., C₁-C₁₂alkoxy), or 1 to 6 carbon atoms(i.e., C₁-C₆ alkoxy). Examples ofsuitable alkoxy groups include, but are not limited to, methoxy (—O—CH₃or —OMe), ethoxy (—OCH₂CH₃ or —OEt), t-butoxy (—O—C(CH₃)₃ or —OtBu) andthe like.

“Haloalkyl” is an alkyl group, as defined above, in which one or morehydrogen atoms of the alkyl group is replaced with a halogen atom. Thealkyl portion of a haloalkyl group can have 1 to 20 carbon atoms (i.e.,C₁-C₂₀ haloalkyl), 1 to 12 carbon atoms(i.e., C₁-C₁₂ haloalkyl), or 1 to6 carbon atoms(i.e., C₁-C₆ alkyl). Examples of suitable haloalkyl groupsinclude, but are not limited to, —CF₃, —CHF₂, —CFH₂, —CH₂CF₃, and thelike. “Alkenyl” is a hydrocarbon containing normal, secondary, tertiaryor cyclic carbon atoms with at least one site of unsaturation, i.e. acarbon-carbon, sp² double bond. For example, an alkenyl group can have 2to 20 carbon atoms (i.e., C₂-C₂₀ alkenyl), 2 to 8 carbon atoms (i.e.,C₂-C₈ alkenyl), or 2 to 6 carbon atoms (i.e., C₂-C₆ alkenyl). Examplesof suitable alkenyl groups include, but are not limited to, ethylene orvinyl (—CH═CH₂), allyl (—CH₂CH═CH₂), cyclopentenyl (—C₅H₇), and5-hexenyl (—CH₂CH₂CH₂CH₂CH═CH₂).

“Alkynyl” is a hydrocarbon containing normal, secondary, tertiary orcyclic carbon atoms with at least one site of unsaturation, i.e. acarbon-carbon, sp triple bond. For example, an alkynyl group can have 2to 20 carbon atoms (i.e., C₂-C₂₀ alkynyl), 2 to 8 carbon atoms (i.e.,C₂-C₈ alkyne,), or 2 to 6 carbon atoms (i.e., C₂-C₆ alkynyl). Examplesof suitable alkynyl groups include, but are not limited to, acetylenic(—C≡CH), propargyl (—CH₂C≡CH), and the like.

“Alkylene” refers to a saturated, branched or straight chain or cyclichydrocarbon radical having two monovalent radical centers derived by theremoval of two hydrogen atoms from the same or two different carbonatoms of a parent alkane. For example, an alkylene group can have 1 to20 carbon atoms, 1 to 10 carbon atoms, or 1 to 6 carbon atoms. Typicalalkylene radicals include, but are not limited to, methylene (—CH₂—),1,1-ethyl (—CH(CH₃)—), 1,2-ethyl (—CH₂CH₂—), 1,1-propyl (—CH(CH₂CH₃)—),1,2-propyl (—CH₂CH(CH₃)—), 1,3-propyl (—CH₂CH₂CH₂—), 1,4-butyl(—CH₂CH₂CH₂CH₂—), and the like.

“Alkenylene” refers to an unsaturated, branched or straight chain orcyclic hydrocarbon radical having two monovalent radical centers derivedby the removal of two hydrogen atoms from the same or two differentcarbon atoms of a parent alkene. For example, and alkenylene group canhave 1 to 20 carbon atoms, 1 to 10 carbon atoms, or 1 to 6 carbon atoms.Typical alkenylene radicals include, but are not limited to,1,2-ethylene (—CH═CH—).

“Alkynylene” refers to an unsaturated, branched or straight chain orcyclic hydrocarbon radical having two monovalent radical centers derivedby the removal of two hydrogen atoms from the same or two differentcarbon atoms of a parent alkyne. For example, an alkynylene group canhave 1 to 20 carbon atoms, 1 to 10 carbon atoms, or 1 to 6 carbon atoms.Typical alkynylene radicals include, but are not limited to, acetylene(—C≡C—), propargyl (—CH₂C≡C—), and 4-pentynyl (—CH₂CH₂CH₂C≡C—).

“Amino” refers generally to a nitrogen radical which can be considered aderivative of ammonia, having the formula —N(X)₂, where each “X” isindependently H, substituted or unsubstituted alkyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,etc. The hybridization of the nitrogen is approximately sp³. Nonlimitingtypes of amino include —NH₂, —N(alkyl)₂, —NH(alkyl), —N(carbocyclyl)₂,—NH(carbocyclyl), —N(heterocyclyl)₂, —NH(heterocyclyl), —N(aryl)₂,—NH(aryl), —N(alkyl)(aryl), —N(alkyl)(heterocyclyl),—N(carbocyclyl)(heterocyclyl), —N(aryl)(heteroaryl),—N(alkyl)(heteroaryl), etc. The term “alkylamino” refers to an aminogroup substituted with at least one alkyl group. Nonlimiting examples ofamino groups include —NH₂, —NH(CH₃), —N(CH₃)₂, —NH(CH₂CH₃), —N(CH₂CH₃)₂,—NH(phenyl), —N(phenyl)₂, —NH(benzyl), —N(benzyl)₂, etc. Substitutedalkylamino refers generally to alkylamino groups, as defined above, inwhich at least one substituted alkyl, as defined herein, is attached tothe amino nitrogen atom. Non-limiting examples of substituted alkylaminoincludes —NH(alkylene-C(O)—OH), —NH(alkylene-C(O)—O-alkyl),—N(alkylene-C(O)—OH)₂, —N(alkylene-C(O)—O-alkyl)₂, etc.

“Aryl” means an aromatic hydrocarbon radical derived by the removal ofone hydrogen atom from a single carbon atom of a parent aromatic ringsystem. For example, an aryl group can have 6 to 20 carbon atoms, 6 to14 carbon atoms, or 6 to 10 carbon atoms. Typical aryl groups include,but are not limited to, radicals derived from benzene (e.g., phenyl),substituted benzene, naphthalene, anthracene, biphenyl, and the like.

“Arylalkyl” refers to an acyclic alkyl radical in which one of thehydrogen atoms bonded to a carbon atom, typically a terminal or sp³carbon atom, is replaced with an aryl radical. Typical arylalkyl groupsinclude, but are not limited to, benzyl, 2-phenylethan-1-yl,naphthylmethyl, 2-naphthylethan-1-yl, naphthobenzyl,2-naphthophenylethan-1-yl and the like. The arylalkyl group can comprise7 to 20 carbon atoms, e.g., the alkyl moiety is 1 to 6 carbon atoms andthe aryl moiety is 6 to 14 carbon atoms.

“Arylalkenyl” refers to an acyclic alkenyl radical in which one of thehydrogen atoms bonded to a carbon atom, typically a terminal or sp³carbon atom, but also an sp² carbon atom, is replaced with an arylradical. The aryl portion of the arylalkenyl can include, for example,any of the aryl groups disclosed herein, and the alkenyl portion of thearylalkenyl can include, for example, any of the alkenyl groupsdisclosed herein. The arylalkenyl group can comprise 8 to 20 carbonatoms, e.g., the alkenyl moiety is 2 to 6 carbon atoms and the arylmoiety is 6 to 14 carbon atoms.

“Arylalkynyl” refers to an acyclic alkynyl radical in which one of thehydrogen atoms bonded to a carbon atom, typically a terminal or sp³carbon atom, but also an sp carbon atom, is replaced with an arylradical. The aryl portion of the arylalkynyl can include, for example,any of the aryl groups disclosed herein, and the alkynyl portion of thearylalkynyl can include, for example, any of the alkynyl groupsdisclosed herein. The arylalkynyl group can comprise 8 to 20 carbonatoms, e.g., the alkynyl moiety is 2 to 6 carbon atoms and the arylmoiety is 6 to 14 carbon atoms.

The term “substituted” in reference to alkyl, alkylene, aryl, arylalkyl,alkoxy, heterocyclyl, heteroaryl, carbocyclyl, etc. , for example,“substituted alkyl”, “substituted alkylene”, “substituted aryl”,“substituted arylalkyl”, “substituted heterocyclyl”, and “substitutedcarbocyclyl” means alkyl, alkylene, aryl, arylalkyl, heterocyclyl,carbocyclyl respectively, in which one or more hydrogen atoms are eachindependently replaced with a non-hydrogen substituent. Typicalsubstituents include, but are not limited to, —X, —R^(b), —O⁻, ═O,—OR^(b), —SR^(b), —S⁻, —NR^(b) ₂, —N⁺R^(b) ₃, ═NR^(b), —CX₃, —CN, —OCN,—SCN, —N═C═O, —NCS, —NO, —NO₂, ═N₂, —N₃, —NHC(═O)R^(b), —OC(═O)R^(b),—NHC(═O)NR —S(═O)₂—, —S(═O)₂OH, —S(═O)₂R^(b), —OS(═O)₂OR^(b),—S(═O)₂NR^(b) ₂, —S(═O)R^(b), —OP(═O)(OR^(b))₂, —P(═O)(OR^(b))₂,—P(═O)(O⁻)₂, —P(═O)(OH)₂, —P(O)(OR^(b))(OR^(b))(O⁻), —C(═O)R^(b),—C(═O)X, —C(S)R^(b), —C(O)OR^(b), —C(O)O⁻, —C(S)OR^(b), —C(O)SR^(b),—C(S)SR^(b), —C(O)NR^(b) ₂, —C(S)NR^(b) ₂, —C(═NR^(b))NR^(b) ₂, whereeach X is independently a halogen: F, Cl, Br, or I; and each R^(b) isindependently H, alkyl, aryl, arylalkyl, a heterocycle, or a protectinggroup or prodrug moiety. Alkylene, alkenylene, and alkynylene groups mayalso be similarly substituted. Unless otherwise indicated, when the term“substituted” is used in conjunction with groups such as arylalkyl,which have two or more moieties capable of substitution, thesubstituents can be attached to the aryl moiety, the alkyl moiety, orboth.

The term “prodrug” as used herein refers to any compound that whenadministered to a biological system generates the drug substance, i.e.,active ingredient, as a result of spontaneous chemical reaction(s),enzyme catalyzed chemical reaction(s), photolysis, and/or metabolicchemical reaction(s). A prodrug is thus a covalently modified analog orlatent form of a therapeutically active compound.

One skilled in the art will recognize that substituents and othermoieties of the compounds of Formula I-IV should be selected in order toprovide a compound which is sufficiently stable to provide apharmaceutically useful compound which can be formulated into anacceptably stable pharmaceutical composition. Compounds of Formula I-IVwhich have such stability are contemplated as falling within the scopeof the present invention.

“Heteroalkyl” refers to an alkyl group where one or more carbon atomshave been replaced with a heteroatom, such as, O, N, or S. For example,if the carbon atom of the alkyl group which is attached to the parentmolecule is replaced with a heteroatom (e.g., O, N, or S) the resultingheteroalkyl groups are, respectively, an alkoxy group (e.g., —OCH₃,etc.), an amine (e.g., —NHCH₃, —N(CH₃)₂, etc.), or a thioalkyl group(e.g., —SCH₃). If a non-terminal carbon atom of the alkyl group which isnot attached to the parent molecule is replaced with a heteroatom (e.g.,O, N, or S) the resulting heteroalkyl groups are, respectively, an alkylether (e.g., —CH₂CH₂—O—CH₃, etc.), an alkyl amine (e.g., —CH₂NHCH₃,—CH₂N(CH₃)_(2,) etc.), or a thioalkyl ether (e.g.,—CH₂—S—CH₃). If aterminal carbon atom of the alkyl group is replaced with a heteroatom(e.g., O, N, or S), the resulting heteroalkyl groups are, respectively,a hydroxyalkyl group (e.g., —CH₂CH₂—OH), an aminoalkyl group (e.g.,—CH₂NH₂), or an alkyl thiol group (e.g., —CH₂CH₂—SH). A heteroalkylgroup can have, for example, 1 to 20 carbon atoms, 1 to 10 carbon atoms,or 1 to 6 carbon atoms. A C₁-C₆ heteroalkyl group means a heteroalkylgroup having 1 to 6 carbon atoms.

“Heterocycle” or “heterocyclyl” as used herein includes by way ofexample and not limitation those heterocycles described in Paquette, LeoA.; Principles of Modern Heterocyclic Chemistry (W.A. Benjamin, NewYork, 1968), particularly Chapters 1, 3, 4, 6, 7, and 9; The Chemistryof Heterocyclic Compounds, A Series of Monographs” (John Wiley & Sons,New York, 1950 to present), in particular Volumes 13, 14, 16, 19, and28; and J. Am. Chem. Soc. (1960) 82:5566. In one specific embodiment ofthe invention “heterocycle” includes a “carbocycle” as defined herein,wherein one or more (e.g. 1, 2, 3, or 4) carbon atoms have been replacedwith a heteroatom (e.g. O, N, or S). The terms “heterocycle” or“heterocyclyl” includes saturated rings, partially unsaturated rings,and aromatic rings (i.e., heteroaromatic rings). Substitutedheterocyclyls include, for example, heterocyclic rings substituted withany of the substituents disclosed herein including carbonyl groups. Anon-limiting example of a carbonyl substituted heterocyclyl is:

Examples of heterocycles include by way of example and not limitationpyridyl, dihydroypyridyl, tetrahydropyridyl (piperidyl), thiazolyl,tetrahydrothiophenyl, sulfur oxidized tetrahydrothiophenyl, pyrimidinyl,furanyl, thienyl, pyrrolyl, pyrazolyl, imidazolyl, tetrazolyl,benzofuranyl, thianaphthalenyl, indolyl, indolenyl, quinolinyl,isoquinolinyl, benzimidazolyl, piperidinyl, 4-piperidonyl, pyrrolidinyl,2-pyrrolidonyl, pyrrolinyl, tetrahydrofuranyl, tetrahydroquinolinyl,tetrahydroisoquinolinyl, decahydroquinolinyl, octahydroisoquinolinyl,azocinyl, triazinyl, 6H-1,2,5-thiadiazinyl, 2H,6H-1,5,2-dithiazinyl,thienyl, thianthrenyl, pyranyl, isobenzofuranyl, chromenyl, xanthenyl,phenoxathinyl, 2H-pyrrolyl, isothiazolyl, isoxazolyl, pyrazinyl,pyridazinyl, indolizinyl, isoindolyl, 3H-indolyl, 1H-indazoly, purinyl,4H-quinolizinyl, phthalazinyl, naphthyridinyl, quinoxalinyl,quinazolinyl, cinnolinyl, pteridinyl, 4aH-carbazolyl, carbazolyl,β-carbolinyl, phenanthridinyl, acridinyl, pyrimidinyl, phenanthrolinyl,phenazinyl, phenothiazinyl, furazanyl, phenoxazinyl, isochromanyl,chromanyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl,piperazinyl, indolinyl, isoindolinyl, quinuclidinyl, morpholinyl,oxazolidinyl, benzotriazolyl, benzisoxazolyl, oxindolyl, benzoxazolinyl,isatinoyl, and bis-tetrahydrofuranyl:

By way of example and not limitation, carbon bonded heterocycles arebonded at position 2, 3, 4, 5, or 6 of a pyridine, position 3, 4, 5, or6 of a pyridazine, position 2, 4, 5, or 6 of a pyrimidine, position 2,3, 5, or 6 of a pyrazine, position 2, 3, 4, or 5 of a furan,tetrahydrofuran, thiofuran, thiophene, pyrrole or tetrahydropyrrole,position 2, 4, or 5 of an oxazole, imidazole or thiazole, position 3, 4,or 5 of an isoxazole, pyrazole, or isothiazole, position 2 or 3 of anaziridine, position 2, 3, or 4 of an azetidine, position 2, 3, 4, 5, 6,7, or 8 of a quinoline or position 1, 3, 4, 5, 6, 7, or 8 of anisoquinoline. Still more typically, carbon bonded heterocycles include2-pyridyl, 3-pyridyl, 4-pyridyl, 5-pyridyl, 6-pyridyl, 3-pyridazinyl,4-pyridazinyl, 5-pyridazinyl, 6-pyridazinyl, 2-pyrimidinyl,4-pyrimidinyl, 5-pyrimidinyl, 6-pyrimidinyl, 2-pyrazinyl, 3-pyrazinyl,5-pyrazinyl, 6-pyrazinyl, 2-thiazolyl, 4-thiazolyl, or 5-thiazolyl.

By way of example and not limitation, nitrogen bonded heterocycles arebonded at position 1 of an aziridine, azetidine, pyrrole, pyrrolidine,2-pyrroline, 3-pyrroline, imidazole, imidazolidine, 2-imidazoline,3-imidazoline, pyrazole, pyrazoline, 2-pyrazoline, 3-pyrazoline,piperidine, piperazine, indole, indoline, 1H-indazole, position 2 of aisoindole, or isoindoline, position 4 of a morpholine, and position 9 ofa carbazole, or β-carboline. Still more typically, nitrogen bondedheterocycles include 1-aziridyl, 1-azetedyl, 1-pyrrolyl, 1-imidazolyl,1-pyrazolyl, and 1-piperidinyl.

“Heterocyclylalkyl” refers to an acyclic alkyl radical in which one ofthe hydrogen atoms bonded to a carbon atom, typically a terminal or sp³carbon atom, is replaced with a heterocyclyl radical (i.e., aheterocyclyl-alkylene-moiety). Typical heterocyclyl alkyl groupsinclude, but are not limited to heterocyclyl-CH₂—,2-(heterocyclyl)ethan-1-yl, and the like, wherein the “heterocyclyl”portion includes any of the heterocyclyl groups described above,including those described in Principles of Modern HeterocyclicChemistry. One skilled in the art will also understand that theheterocyclyl group can be attached to the alkyl portion of theheterocyclyl alkyl by means of a carbon-carbon bond or acarbon-heteroatom bond, with the proviso that the resulting group ischemically stable. The heterocyclyl alkyl group comprises 3 to 20 carbonatoms, e.g., the alkyl portion of the arylalkyl group is 1 to 6 carbonatoms and the heterocyclyl moiety is 2 to 14 carbon atoms. Examples ofheterocyclylalkyls include by way of example and not limitation5-membered sulfur, oxygen, and/or nitrogen containing heterocycles suchas thiazolylmethyl, 2-thiazolylethan-1-yl, imidazolylmethyl,oxazolylmethyl, thiadiazolylmethyl, etc., 6-membered sulfur, oxygen,and/or nitrogen containing heterocycles such as piperidinylmethyl,piperazinylmethyl, morpholinylmethyl, pyridinylmethyl, pyridizylmethyl,pyrimidylmethyl, pyrazinylmethyl, etc.

“Heterocyclylalkenyl” refers to an acyclic alkenyl radical in which oneof the hydrogen atoms bonded to a carbon atom, typically a terminal orsp³ carbon atom, but also a sp² carbon atom, is replaced with aheterocyclyl radical (i.e., a heterocyclyl-alkenylene-moiety). Theheterocyclyl portion of the heterocyclyl alkenyl group includes any ofthe heterocyclyl groups described herein, including those described inPrinciples of Modern Heterocyclic Chemistry, and the alkenyl portion ofthe heterocyclyl alkenyl group includes any of the alkenyl groupsdisclosed herein. One skilled in the art will also understand that theheterocyclyl group can be attached to the alkenyl portion of theheterocyclyl alkenyl by means of a carbon-carbon bond or acarbon-heteroatom bond, with the proviso that the resulting group ischemically stable. The heterocyclyl alkenyl group comprises 4 to 20carbon atoms, e.g., the alkenyl portion of the heterocyclyl alkenylgroup is 2 to 6 carbon atoms and the heterocyclyl moiety is 2 to 14carbon atoms.

“Heterocyclylalkynyl” refers to an acyclic alkynyl radical in which oneof the hydrogen atoms bonded to a carbon atom, typically a terminal orsp³ carbon atom, but also an sp carbon atom, is replaced with aheterocyclyl radical (i.e., a heterocyclyl-alkynylene-moiety). Theheterocyclyl portion of the heterocyclyl alkynyl group includes any ofthe heterocyclyl groups described herein, including those described inPrinciples of Modern Heterocyclic Chemistry, and the alkynyl portion ofthe heterocyclyl alkynyl group includes any of the alkynyl groupsdisclosed herein. One skilled in the art will also understand that theheterocyclyl group can be attached to the alkynyl portion of theheterocyclyl alkynyl by means of a carbon-carbon bond or acarbon-heteroatom bond, with the proviso that the resulting group ischemically stable. The heterocyclyl alkynyl group comprises 4 to 20carbon atoms, e.g., the alkynyl portion of the heterocyclyl alkynylgroup is 2 to 6 carbon atoms and the heterocyclyl moiety is 2 to 14carbon atoms.

“Heteroaryl” refers to an aromatic heterocyclyl having at least oneheteroatom in the ring. Non-limiting examples of suitable heteroatomswhich can be included in the aromatic ring include oxygen, sulfur, andnitrogen. Non-limiting examples of heteroaryl rings include all of thosearomatic rings listed in the definition of “heterocyclyl”, includingpyridinyl, pyrrolyl, oxazolyl, indolyl, isoindolyl, purinyl, furanyl,thienyl, benzofuranyl, benzothiophenyl, carbazolyl, imidazolyl,thiazolyl, isoxazolyl, pyrazolyl, isothiazolyl, quinolyl, isoquinolyl,pyridazyl, pyrimidyl, pyrazyl, etc.

“Carbocycle” or “carbocyclyl” refers to a saturated (i.e., cycloalkyl),partially unsaturated (e.g., cycloakenyl, cycloalkadienyl, etc.) oraromatic ring having 3 to 7 carbon atoms as a monocycle, 7 to 12 carbonatoms as a bicycle, and up to about 20 carbon atoms as a polycycle.Monocyclic carbocycles have 3 to 7 ring atoms, still more typically 5 or6 ring atoms. Bicyclic carbocycles have 7 to 12 ring atoms, e.g.,arranged as a bicyclo [4,5], [5,5], [5,6] or [6,6] system, or 9 or 10ring atoms arranged as a bicyclo [5,6] or [6,6] system, or spiro-fusedrings. Non-limiting examples of monocyclic carbocycles includecyclopropyl, cyclobutyl, cyclopentyl, 1-cyclopent-1-enyl,1-cyclopent-2-enyl, 1-cyclopent-3-enyl, cyclohexyl, 1-cyclohex-1-enyl,1-cyclohex-2-enyl, 1-cyclohex-3-enyl, and phenyl. Non-limiting examplesof bicyclo carbocycles includes naphthyl, tetrahydronapthalene, anddecaline.

“Carbocyclylalkyl” refers to to an acyclic akyl radical in which one ofthe hydrogen atoms bonded to a carbon atom is replaced with acarbocyclyl radical as described herein. Typical, but non-limiting,examples of carbocyclylalkyl groups include cyclopropylmethyl,cyclopropylethyl, cyclobutylmethyl, cyclopentylmethyl andcyclohexylmethyl.

“Arylheteroalkyl” refers to a heteroalkyl as defined herein, in which ahydrogen atom (which may be attached either to a carbon atom or aheteroatom) has been replaced with an aryl group as defined herein. Thearyl groups may be bonded to a carbon atom of the heteroalkyl group, orto a heteroatom of the heteroalkyl group, provided that the resultingarylheteroalkyl group provides a chemically stable moiety. For example,an arylheteroalkyl group can have the general formulae -alkylene-O-aryl,-alkylene-O-alkylene-aryl, -alkylene-NH-aryl,-alkylene-NH-alkylene-aryl, -alkylene-S-aryl, -alkylene-S-alkylene-aryl,etc. In addition, any of the alkylene moieties in the general formulaeabove can be further substituted with any of the substituents defined orexemplified herein.

“Heteroarylalkyl” refers to an alkyl group, as defined herein, in whicha hydrogen atom has been replaced with a heteroaryl group as definedherein. Non-limiting examples of heteroaryl alkyl include—CH₂-pyridinyl, —CH₂-pyrrolyl, —CH₂-oxazolyl, —CH₂-indolyl,—CH₂-isoindolyl, —CH₂-purinyl, —CH₂-furanyl, —CH₂-thienyl,—CH₂-benzofuranyl, —CH₂-benzothiophenyl, —CH₂-carbazolyl,—CH₂-imidazolyl, —CH₂-thiazolyl, —CH₂-isoxazolyl, —CH₂-pyrazolyl,—CH₂-isothiazolyl, —CH₂-quinolyl, —CH₂-isoquinolyl, —CH₂-pyridazyl,—CH₂-pyrimidyl, —CH₂-pyrazyl, —CH(CH₃)-pyridinyl, —CH(CH₃)-pyrrolyl,—CH(CH₃)-oxazolyl, —CH(CH₃)-indolyl, —CH(CH₃)-isoindolyl,—CH(CH₃)-purinyl, —CH(CH₃)-furanyl, —CH(CH₃)-thienyl,—CH(CH₃)-benzofuranyl, —CH(CH₃)-benzothiophenyl, —CH(CH₃)-carbazolyl,—CH(CH₃)-imidazolyl, —CH(CH₃)-thiazolyl, —CH(CH₃)-isoxazolyl,—CH(CH₃)-pyrazolyl, —CH(CH₃)-isothiazolyl, —CH(CH₃)-quinolyl,—CH(CH₃)-isoquinolyl, —CH(CH₃)-pyridazyl, —CH(CH₃)-pyrimidyl,—CH(CH₃)-pyrazyl, etc.

The term “optionally substituted” in reference to a particular moiety ofthe compound of Formula I-III (e.g., an optionally substituted arylgroup) refers to a moiety wherein all substituents are hydrogen orwherein one or more of the hydrogens of the moiety may be replaced bysubstituents such as those listed under the definition of “substituted”.

The term “optionally replaced” in reference to a particular moiety ofthe compound of Formula I-III (e.g., the carbon atoms of said(C₁-C₈)alkyl may be optionally replaced by —O—, —S—, or —NR^(a)—) meansthat one or more of the methylene groups of the (C₁-C₈)alkyl may bereplaced by 0, 1, 2, or more of the groups specified (e.g., —O—, —S—, or—NR^(a)—).

The term “non-terminal carbon atom(s)” in reference to an alkyl,alkenyl, alkynyl, alkylene, alkenylene, or alkynylene moiety refers tothe carbon atoms in the moiety that intervene between the first carbonatom of the moiety and the last carbon atom in the moiety. Therefore, byway of example and not limitation, in the alkyl moiety—CH₂(C*)H₂(C*)H₂CH₃ or alkylene moiety —CH₂(C*)H₂(C*)H₂CH₂— the C* atomswould be considered to be the non-terminal carbon atoms.

Certain Y and Y¹ alternatives are nitrogen oxides such as ⁺N(O)(R) or⁺N(O)(OR). These nitrogen oxides, as shown here attached to a carbonatom, can also be represented by charge separated groups such as

respectively, and are intended to be equivalent to the aforementionedrepresentations for the purposes of describing this invention.

“Linker” or “link” means a chemical moiety comprising a covalent bond ora chain of atoms. Linkers include repeating units of alkyloxy (e.g.polyethyleneoxy, PEG, polymethyleneoxy) and alkylamino (e.g.polyethyleneamino, Jeffamine™); and diacid ester and amides includingsuccinate, succinamide, diglycolate, malonate, and caproamide.

The terms such as “oxygen-linked”, “nitrogen-linked”, “carbon-linked”,“sulfur-linked”, or “phosphorous-linked” mean that if a bond between twomoieties can be formed by using more than one type of atom in a moiety,then the bond formed between the moieties is through the atom specified.For example, a nitrogen-linked amino acid would be bonded through anitrogen atom of the amino acid rather than through an oxygen or carbonatom of the amino acid.

In some embodiments of the compounds of Formula I-IV, one or more of W¹or W² are independently a radical of a nitrogen-linked naturallyoccurring α-amino acid ester. Examples of naturally occurring aminoacids include isoleucine, leucine, lysine, methionine, phenylalanine,threonine, tryptophan, valine, alanine, asparagine, aspartic acid,cysteine, glutamic acid, glutamine, glycine, proline, selenocysteine,serine, tyrosine, arginine, histidine, ornithine and taurine. The estersof these amino acids comprise any of those described for thesubstitutent R, particularly those in which R is optionally substituted(C₁-C₈)alkyl.

The term “purine” or “pyrimidine” base comprises, but is not limited to,adenine, N⁶-alkylpurines, N⁶-acylpurines (wherein acyl is C(O)(alkyl,aryl, alkylaryl, or arylalkyl), N⁶-benzylpurine, N⁶-halopurine,N⁶-vinylpurine, N⁶-acetylenic purine, N⁶-acyl purine, N⁶-hydroxyalkylpurine, N⁶-allylaminopurine, N⁶-thioallyl purine, N²-alkylpurines,N²-alkyl-6-thiopurines, thymine, cytosine, 5-fluorocytosine,5-methylcytosine, 6-azapyrimidine, including 6-azacytosine, 2- and/or4-mercaptopyrmidine, uracil, 5-halouracil, including 5-fluorouracil,C⁵-alkylpyrimidines, C⁵-benzylpyrimidines, C⁵-halopyrimidines,C⁵-vinylpyrimidine, C⁵-acetylenic pyrimidine, C⁵-acyl pyrimidine,C⁵-hydroxyalkyl purine, C⁵-amidopyrimidine, C⁵-cyanopyrimidine,C⁵-5-iodopyrimidine, C⁶-iodo-pyrimidine, C⁵-Br-vinyl pyrimidine,C⁶-Br-vinyl pyriniidine, C⁵-nitropyrimidine, C⁵-amino-pyrimidine,N²-alkylpurines, N²-alkyl-6-thiopurines, 5-azacytidinyl, 5-azauracilyl,triazolopyridinyl, imidazolopyridinyl, pyrrolopyrimidinyl, andpyrazolopyrimidinyl. Purine bases include, but are not limited to,guanine, adenine, hypoxanthine, 2,6-diaminopurine, and 6-chloropurine.The purine and pyrimidine bases of Formula I-III are linked to theribose sugar, or analog thereof, through a nitrogen atom of the base.Functional oxygen and nitrogen groups on the base can be protected asnecessary or desired. Suitable protecting groups are well known to thoseskilled in the art, and include trimethylsilyl, dimethylhexylsilyl,t-butyldimethylsilyl, and t-butyldiphenylsilyl, trityl, alkyl groups,and acyl groups such as acetyl and propionyl, methanesulfonyl, andp-toluenesulfonyl.

Unless otherwise specified, the carbon atoms of the compounds of FormulaI-IV are intended to have a valence of four. In some chemical structurerepresentations where carbon atoms do not have a sufficient number ofvariables attached to produce a valence of four, the remaining carbonsubstitutents needed to provide a valence of four should be assumed tobe hydrogen. For example,

has the same meaning as

“Protecting group” refers to a moiety of a compound that masks or altersthe properties of a functional group or the properties of the compoundas a whole. The chemical substructure of a protecting group varieswidely. One function of a protecting group is to serve as anintermediate in the synthesis of the parental drug substance. Chemicalprotecting groups and strategies for protection/deprotection are wellknown in the art. See: “Protective Groups in Organic Chemistry”,Theodora W. Greene (John Wiley & Sons, Inc., New York, 1991. Protectinggroups are often utilized to mask the reactivity of certain functionalgroups, to assist in the efficiency of desired chemical reactions, e.g.making and breaking chemical bonds in an ordered and planned fashion.Protection of functional groups of a compound alters other physicalproperties besides the reactivity of the protected functional group,such as the polarity, lipophilicity (hydrophobicity), and otherproperties which can be measured by common analytical tools. Chemicallyprotected intermediates may themselves be biologically active orinactive.

Protected compounds may also exhibit altered, and in some cases,optimized properties in vitro and in vivo, such as passage throughcellular membranes and resistance to enzymatic degradation orsequestration. In this role, protected compounds with intendedtherapeutic effects may be referred to as prodrugs. Another function ofa protecting group is to convert the parental drug into a prodrug,whereby the parental drug is released upon conversion of the prodrug invivo. Because active prodrugs may be absorbed more effectively than theparental drug, prodrugs may possess greater potency in vivo than theparental drug. Protecting groups are removed either in vitro, in theinstance of chemical intermediates, or in vivo, in the case of prodrugs.With chemical intermediates, it is not particularly important that theresulting products after deprotection, e.g. alcohols, be physiologicallyacceptable, although in general it is more desirable if the products arepharmacologically innocuous.

“Prodrug moiety” means a labile functional group which separates fromthe active inhibitory compound during metabolism, systemically, inside acell, by hydrolysis, enzymatic cleavage, or by some other process(Bundgaard, Hans, “Design and Application of Prodrugs” in Textbook ofDrug Design and Development (1991), P. Krogsgaard-Larsen and H.Bundgaard, Eds. Harwood Academic Publishers, pp. 113-191). Enzymes whichare capable of an enzymatic activation mechanism with the phosphonateprodrug compounds of the invention include, but are not limited to,amidases, esterases, microbial enzymes, phospholipases, cholinesterases,and phosphases. Prodrug moieties can serve to enhance solubility,absorption and lipophilicity to optimize drug delivery, bioavailabilityand efficacy.

A prodrug moiety may include an active metabolite or drug itself.

Exemplary prodrug moieties include the hydrolytically sensitive orlabile acyloxymethyl esters —CH₂OC(═O)R³⁰ and acyloxymethyl carbonates—CH₂OC(═O)OR³⁰ where R³⁰ is C₁-C₆ alkyl, C₁-C₆ substituted alkyl, C₆-C₂₀aryl or C₆-C₂₀ substituted aryl. The acyloxyalkyl ester was used as aprodrug strategy for carboxylic acids and then applied to phosphates andphosphonates by Farquhar et al (1983) J. Pharm. Sci. 72: 324; also U.S.Pat. Nos. 4,816,570, 4,968,788, 5,663,159 and 5,792,756. In certaincompounds of the invention, a prodrug moiety is part of a phosphategroup. The acyloxyalkyl ester may be used to deliver phosphoric acidsacross cell membranes and to enhance oral bioavailability. A closevariant of the acyloxyalkyl ester, the alkoxycarbonyloxyalkyl ester(carbonate), may also enhance oral bioavailability as a prodrug moietyin the compounds of the combinations of the invention. An exemplaryacyloxymethyl ester is pivaloyloxymethoxy, (POM) —CH₂OC(═O)C(CH₃)₃. Anexemplary acyloxymethyl carbonate prodrug moiety ispivaloyloxymethylcarbonate (POC) —CH₂OC(═O)OC(CH₃)₃.

The phosphate group may be a phosphate prodrug moiety. The prodrugmoiety may be sensitive to hydrolysis, such as, but not limited to thosecomprising a pivaloyloxymethyl carbonate (POC) or POM group.Alternatively, the prodrug moiety may be sensitive to enzymaticpotentiated cleavage, such as a lactate ester or a phosphonamidate-estergroup.

Aryl esters of phosphorus groups, especially phenyl esters, are reportedto enhance oral bioavailability (DeLambert et al (1994) J. Med. Chem.37: 498). Phenyl esters containing a carboxylic ester ortho to thephosphate have also been described (Khamnei and Torrence, (1996) J. Med.Chem. 39:4109-4115). Benzyl esters are reported to generate the parentphosphonic acid. In some cases, substituents at the ortho- orpara-position may accelerate the hydrolysis. Benzyl analogs with anacylated phenol or an alkylated phenol may generate the phenoliccompound through the action of enzymes, e.g. esterases, oxidases, etc.,which in turn undergoes cleavage at the benzylic C—O bond to generatethe phosphoric acid and the quinone methide intermediate. Examples ofthis class of prodrugs are described by Mitchell et al (1992) J. Chem.Soc. Perkin Trans. 12345; Brook et al WO 91/19721. Still other benzylicprodrugs have been described containing a carboxylic ester-containinggroup attached to the benzylic methylene (Glazier et al WO 91/19721).Thio-containing prodrugs are reported to be useful for the intracellulardelivery of phosphonate drugs. These proesters contain an ethylthiogroup in which the thiol group is either esterified with an acyl groupor combined with another thiol group to form a disulfide.Deesterification or reduction of the disulfide generates the free thiointermediate which subsequently breaks down to the phosphoric acid andepisulfide (Puech et al (1993) Antiviral Res., 22: 155-174; Benzaria etal (1996) J. Med. Chem. 39: 4958). Cyclic phosphonate esters have alsobeen described as prodrugs of phosphorus-containing compounds (Erion etal, U.S. Pat. No. 6312662).

It is to be noted that all enantiomers, diastereomers, and racemicmixtures, tautomers, polymorphs, pseudopolymorphs of compounds withinthe scope of Formula I-IV and pharmaceutically acceptable salts thereofare embraced by the present invention. All mixtures of such enantiomersand diastereomers are within the scope of the present invention.

A compound of Formula I-IV and its pharmaceutically acceptable salts mayexist as different polymorphs or pseudopolymorphs. As used herein,crystalline polymorphism means the ability of a crystalline compound toexist in different crystal structures. The crystalline polymorphism mayresult from differences in crystal packing (packing polymorphism) ordifferences in packing between different conformers of the same molecule(conformational polymorphism). As used herein, crystallinepseudopolymorphism means the ability of a hydrate or solvate of acompound to exist in different crystal structures. The pseudopolymorphsof the instant invention may exist due to differences in crystal packing(packing pseudopolymorphism) or due to differences in packing betweendifferent conformers of the same molecule (conformationalpseudopolymorphism). The instant invention comprises all polymorphs andpseudopolymorphs of the compounds of Formula I-III and theirpharmaceutically acceptable salts.

A compound of Formula I-IV and its pharmaceutically acceptable salts mayalso exist as an amorphous solid. As used herein, an amorphous solid isa solid in which there is no long-range order of the positions of theatoms in the solid. This definition applies as well when the crystalsize is two nanometers or less. Additives, including solvents, may beused to create the amorphous forms of the instant invention. The instantinvention comprises all amorphous forms of the compounds of Formula I-IVand their pharmaceutically acceptable salts.

Selected substituents comprising the compounds of Formula I-IV arepresent to a recursive degree. In this context, “recursive substituent”means that a substituent may recite another instance of itself. Becauseof the recursive nature of such substituents, theoretically, a largenumber of compounds may be present in any given embodiment. For example,R^(x) comprises a R^(y) substituent. R^(y) can be R. R can be W³. W³ canbe W⁴ and W⁴ can be R or comprise substituents comprising R^(y). One ofordinary skill in the art of medicinal chemistry understands that thetotal number of such substituents is reasonably limited by the desiredproperties of the compound intended. Such properties include, by way ofexample and not limitation, physical properties such as molecularweight, solubility or log P, application properties such as activityagainst the intended target, and practical properties such as ease ofsynthesis.

By way of example and not limitation, W³ and R^(y) are recursivesubstituents in certain embodiments. Typically, each recursivesubstituent can independently occur 20, 19, 18, 17, 16, 15, 14, 13, 12,11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, or 0, times in a given embodiment.More typically, each recursive substituent can independently occur 12 orfewer times in a given embodiment. Even more typically, each recursivesubstituent can independently occur 3 or fewer times in a givenembodiment. For example, W³ will occur 0 to 8 times, R^(y) will occur 0to 6 times in a given embodiment. Even more typically, W³ will occur 0to 6 times and R^(y) will occur 0 to 4 times in a given embodiment.

Recursive substituents are an intended aspect of the invention. One ofordinary skill in the art of medicinal chemistry understands theversatility of such substituents. To the degree that recursivesubstituents are present in an embodiment of the invention, the totalnumber will be determined as set forth above.

The modifier “about” used in connection with a quantity is inclusive ofthe stated value and has the meaning dictated by the context (e.g.,includes the degree of error associated with measurement of theparticular quantity).

The term “treating”, as used herein, unless otherwise indicated, meansreversing, alleviating, inhibiting the progress of, or preventing thedisorder or condition to which such term applies, or one or moresymptoms of such disorder or condition. The term “treatment”, as usedherein, refers to the act of treating, as “treating” is definedimmediately above.

The term “therapeutically effective amount”, as used herein, is theamount of compound of Formula I-IV present in a composition describedherein that is needed to provide a desired level of drug in thesecretions and tissues of the airways and lungs, or alternatively, inthe bloodstream of a subject to be treated to give an anticipatedphysiological response or desired biological effect when such acomposition is administered by the chosen route of administration. Theprecise amount will depend upon numerous factors, for example theparticular compound of Formula I-IV, the specific activity of thecomposition, the delivery device employed, the physical characteristicsof the composition, its intended use, as well as patient considerationssuch as severity of the disease state, patient cooperation, etc., andcan readily be determined by one skilled in the art based upon theinformation provided herein.

The term “normal saline” means a water solution containing 0.9% (w/v)NaCl.

The term “hypertonic saline” means a water solution containing greaterthan 0.9% (w/v) NaCl. For example, 3% hypertonic saline would contain 3%(w/v) NaCl.

The compounds of the Formula I-IV may comprise a phosphate group as R⁷,which may be a prodrug moiety

wherein each Y or Y¹ is, independently, O, S, NR, ⁺N(O)(R), N(OR),⁺N(O)(OR), or N—NR₂; W¹ and W², when taken together, are—Y³(C(R^(y))₂)₃Y³—; or one of W¹ or W² together with either R³or R⁴ is—Y³— and the other of W¹ or W² is Formula Ia; or W¹ and W² are each,independently, a group of Formula Ia:

wherein:

each Y² is independently a bond, O, CR₂, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR),N—NR₂, S, S—S, S(O), or S(O)₂;

each Y³ is independently O, S, or NR;

M2 is 0, 1 or 2;

each R^(y) is independently H, F, Cl, Br, I, OH, R, —C(═Y¹)R, —C(═Y¹)OR,—C(═Y¹)N(R)₂, —N(R)₂, —⁺N(R)₃, —SR, —S(O)R, —S(O)₂R, —S(O)(OR),—S(O)₂(OR), —OC(═Y¹)R, —OC(═Y¹)OR, —OC(═Y¹)(N(R)₂), —SC(═Y¹)R,—SC(═Y¹)OR, —SC(═Y¹)(N(R)₂), —N(R)C(═Y¹)R, —N(R)C(═Y¹)OR, or—N(R)C(═Y¹)N(R)₂, —SO₂NR₂, —CN, —N₃, —NO₂, —OR, a protecting group orW³; or when taken together, two R^(y) on the same carbon atom form acarbocyclic ring of 3 to 7 carbon atoms;

each R^(x) is independently R^(y), a protecting group, or the formula:

wherein:

M1a, M1c, and M1d are independently 0 or 1;

M12c is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12;

each R is H, halogen, (C₁-C₈) alkyl, (C₁-C₈) substituted alkyl, (C₂-C₈)alkenyl, (C₂-C₈) substituted alkenyl, (C₂-C₈) alkynyl, (C₂-C₈)substituted alkynyl, C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₂-C₂₀heterocycle, C₂-C₂₀ substituted heterocyclyl, arylalkyl, substitutedarylalkyl or a protecting group;

W³ is W⁴ or W⁵; W⁴ is R, —C(Y¹)R^(y), —C(Y¹)W⁵, —SO₂R^(y), or —SO₂W⁵;and W⁵ is a carbocycle or a heterocycle wherein W⁵ is independentlysubstituted with 0 to 3 R^(y) groups.

W⁵ carbocycles and W⁵ heterocycles may be independently substituted with0 to 3 R^(y) groups. W⁵ may be a saturated, unsaturated or aromatic ringcomprising a mono- or bicyclic carbocycle or heterocycle. W⁵ may have 3to 10 ring atoms, e.g., 3 to 7 ring atoms. The W⁵ rings are saturatedwhen containing 3 ring atoms, saturated or mono-unsaturated whencontaining 4 ring atoms, saturated, or mono- or di-unsaturated whencontaining 5 ring atoms, and saturated, mono- or di-unsaturated, oraromatic when containing 6 ring atoms.

A W⁵ heterocycle may be a monocycle having 3 to 7 ring members (2 to 6carbon atoms and 1 to 3 heteroatoms selected from N, O, P, and S) or abicycle having 7 to 10 ring members (4 to 9 carbon atoms and 1 to 3heteroatoms selected from N, O, P, and S). W⁵ heterocyclic monocyclesmay have 3 to 6 ring atoms (2 to 5 carbon atoms and 1 to 2 heteroatomsselected from N, O, and S); or 5 or 6 ring atoms (3 to 5 carbon atomsand 1 to 2 heteroatoms selected from N and S). W⁵ heterocyclic bicycleshave 7 to 10 ring atoms (6 to 9 carbon atoms and 1 to 2 heteroatomsselected from N, O, and S) arranged as a bicyclo [4,5], [5,5], [5,6], or[6,6] system; or 9 to 10 ring atoms (8 to 9 carbon atoms and 1 to 2hetero atoms selected from N and S) arranged as a bicyclo [5,6] or [6,6]system. The W⁵ heterocycle may be bonded to Y² through a carbon,nitrogen, sulfur or other atom by a stable covalent bond.

W⁵ heterocycles include for example, pyridyl, dihydropyridyl isomers,piperidine, pyridazinyl, pyrimidinyl, pyrazinyl, s-triazinyl, oxazolyl,imidazolyl, thiazolyl, isoxazolyl, pyrazolyl, isothiazolyl, furanyl,thiofuranyl, thienyl, and pyrrolyl. W⁵ also includes, but is not limitedto, examples such as:

W⁵ carbocycles and heterocycles may be independently substituted with 0to 3 R groups, as defined above. For example, substituted W⁵ carbocyclesinclude:

Examples of substituted phenyl carbocycles include:

Embodiments of

of Formula I-IV compounds include substructures such as:

wherein each Y^(2b) is, independently, O or N(R). In another aspect ofthis embodiment, each Y^(2b) is O and each R^(x) is independently:

wherein M12c is 1, 2 or 3 and each Y² is independently a bond, O, CR₂,or S. In another aspect of this embodiment, one Y^(2b)—R^(x) is NH(R)and the other Y^(2b)—R^(x) is O—R^(x) wherein R^(x) is:

wherein M12c is 2. In another aspect of this embodiment, each Y^(2b) isO and each R^(x) is independently:

wherein M12c is 2. In another aspect of this embodiment, each Y^(2b) isO and each R^(x) is independently:

wherein M12c is 1 and Y² is a bond, O, or CR₂.

Other embodiments of

of Formulas I-IV compounds include substructures such as:

wherein each Y³ is, independently, O or N(R). In another aspect of thisembodiment, each Y³ is O. In another aspect of this embodiment, thesubstructure is:

wherein R^(y) is W⁵ as defined herein.

Another embodiment of

of Formula I-IV includes the substructures:

wherein each Y^(2c) is, independently, O, N(R^(y)) or S.

Another embodiment of

of Formula I-IV compounds includes the substructures wherein one of W¹or W² together with either R³ or R⁴ is —Y³— and the other of W¹ or W² isFormula Ia. Such an embodiment is represented by a compound of FormulaIb selected from:

In another aspect of the embodiment of Formula lb, each Y and Y³ is O.In another aspect of the embodiment of Formula lb, W¹ or W² isY^(2b)—R^(x); each Y, Y³ and Y^(2b) is O and R^(x) is:

wherein M12c is 1, 2 or 3 and each Y² is independently a bond, O, CR₂,or S. In another aspect of the embodiment of Formula lb, W¹ or W² isY^(2b)—R^(x); each Y, Y³ and Y^(2b) is O and R^(x) is:

wherein M12c is 2. In another aspect of the embodiment of Formula lb, W¹or W² is Y^(2b)—R^(x); each Y, Y³ and Y^(2b) is O and R^(x) is:

wherein M12c is 1 and Y² is a bond, O, or CR₂.

Another embodiment of

of Formula I-IV compounds includes a substructure:

wherein W⁵ is a carbocycle such as phenyl or substituted phenyl. Inanother aspect of this embodiment, the substructure is:

wherein Y^(2b) is O or N(R) and the phenyl carbocycle is substitutedwith 0 to 3 R groups. In another aspect of this embodiment of thesubstructure, R^(x) is:

wherein M12c is 1, 2 or 3 and each Y² is independently a bond, O, CR₂,or S.

Another embodiment of

of Formula I-IV includes substructures:

The chiral carbon of the amino acid and lactate moieties may be eitherthe R or S configuration or the racemic mixture.

Another embodiment of

of Formula I-IV is substructure

wherein each Y² is, independently, —O— or —NH—. In another aspect ofthis embodiment, R^(y) is (C₁-C₈) alkyl, (C₁-C₈) substituted alkyl,(C₂-C₈) alkenyl, (C₂-C₈) substituted alkenyl, (C₂-C₈) alkynyl or (C₂-C₈)substituted alkynyl. In another aspect of this embodiment, R^(y) is(C₁-C₈) alkyl, (C₁-C₈) substituted alkyl, (C₂-C₈) alkenyl, (C₂-C₈)substituted alkenyl, (C₂-C₈) alkynyl or (C₂-C₈) substituted alkynyl; andR is CH₃. In another aspect of this embodiment, R^(y) is (C₁-C₈) alkyl,(C₁-C₈) substituted alkyl, (C₂-C₈) alkenyl, (C₂-C₈) substituted alkenyl,(C₂-C₈) alkynyl or (C₂-C₈) substituted alkynyl; R is CH₃; and each Y² is—NH—. In another aspect of this embodiment, W¹ and W² are,independently, nitrogen-linked, naturally occurring amino acids ornaturally occurring amino acid esters. In another aspect of thisembodiment, W¹ and W² are, independently, naturally-occurring 2-hydroxycarboxylic acids or naturally-occurring 2-hydroxy carboxylic acid esterswherein the acid or ester is linked to P through the 2-hydroxy group.

Another embodiment of

of Formula I-IV is substructure:

In one aspect of this embodiment, each R^(x) is, independently, (C₁-C₈)alkyl. In another aspect of this embodiment, each R^(x) is,independently, C₆-C₂₀ aryl or C₆-C₂₀ substituted aryl.

In a preferred embodiment,

is selected from

Another embodiment of

of Formulas I-IV is substructure

wherein W¹ and W² are independently selected from one of the formulas inTables 20.1-20.37 and Table 30.1 below. The variables used in Tables20.1-20.37 (e.g., W²³, R²¹, etc.) pertain only to Tables 20.1-20.37,unless otherwise indicated.

The variables used in Tables 20.1 to 20.37 have the followingdefinitions:

each R²¹ is independently H or (C₁-C₈)alkyl;

each R²² is independently H, R²¹, R²³ or R²⁴ wherein each R²⁴ isindependently substituted with 0 to 3 R²³;

each R²³ is independently R^(23a), R^(23b), R^(23c) or R^(23d), providedthat when R²³ is bound to a heteroatom, then R²³ is R^(23c) or R^(23d);

each R^(23a) is independently F, Cl, Br, I, —CN, N₃ or —NO₂;

each R^(23b) is independently Y²¹;

each R^(23c) is independently —R^(2x), —N(R^(2x))(R^(2x)), —SR^(2x),—S(O)R^(2x), —S(O)₂R^(2x), —S(O)(OR^(2x)), —S(O)₂(OR^(2x)),—OC(═Y²¹)R^(2x), —OC(═Y²¹)OR^(2x), —OC(═Y²¹)(N(R^(2x))(R^(2x))),—SC(═Y²¹)R^(2x), —SC(═Y²¹)OR^(2x), —SC(═Y²¹)(N(R^(2x))(R^(2x))),—N(R^(2x))(C(═Y²¹)R^(2x), —N(R^(2x))C(═Y²¹)OR^(2x), or—N(R^(2x))C(═Y²¹)(N(R^(2x))(R^(2x)));

each R^(23d) is independently —C(═Y²¹)R^(2x), —C(═Y²¹)OR^(2x) or—C(═Y²¹)(N(R^(2x))(R^(2x)));

each R^(2x) is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl,(C₂-C₈)alkynyl, aryl, heteroaryl; or two R^(2x) taken together with anitrogen to which they are both attached form a 3 to 7 memberedheterocyclic ring wherein any one carbon atom of said heterocyclic ringcan optionally be replaced with —O—, —S— or —NR²¹—; and wherein one ormore of the non-terminal carbon atoms of each said (C₁-C₈)alkyl may beoptionally replaced with —O—, —S— or —NR²¹—;

each R²⁴ is independently (C₁-C₈)alkyl, (C₂-C₈)alkenyl, or(C₂-C₈)alkynyl;

each R²⁵ is independently R²⁴ wherein each R²⁴ is substituted with 0 to3 R²³ groups;

each R^(25a) is independently (C₁-C₈)alkylene, (C₂-C₈)alkenylene, or(C₂-C₈)alkynylene any one of which said (C₁-C₈)alkylene,(C₂-C₈)alkenylene, or (C₂-C₈)alkynylene is substituted with 0-3 R²³groups;

each W²³ is independently W²⁴ or W²⁵;

each W²⁴ is independently R²⁵, —C(═Y²¹)R²⁵, —C(═Y²¹)W²⁵, —SO₂R²⁵,—SO₂W²⁵;

each W²⁵ is independently carbocycle or heterocycle wherein W²⁵ isindependently substituted with 0 to 3 R²² groups; and

each Y²¹ is independently O or S.

TABLE 20.1

1

2

3

4

5

6

7

8

TABLE 20.2

9

10

11

TABLE 20.3

12

13

14

15

16

17

18

19

TABLE 20.4

20

21

22

TABLE 20.5

23

24

25

26

27

28

29

30

TABLE 20.6

31

32

33

TABLE 20.7

34

35

36

37

38

39

40

41

TABLE 20.8

42

43

44

45

46

47

48

49

TABLE 20.9

50

51

52

53

54

55

56

57

TABLE 20.10

58

59

60

TABLE 20.11

61

62

63

64

65

66

67

68

TABLE 20.12

69

70

71

TABLE 20.13

72

73

74

75

76

77

78

79

TABLE 20.14

80

81

82

TABLE 20.15

83

84

85

86

87

88

89

90

TABLE 20.16

91

92

93

94

95

96

97

98

TABLE 20.17

99

100

101

102

103

104

105

106

TABLE 20.18

107

108

109

TABLE 20.19

110

111

112

113

114

115

116

117

TABLE 20.20

118

119

120

TABLE 20.21

121

122

123

124

125

126

127

128

TABLE 20.22

129

130

131

TABLE 20.23

132

133

134

135

136

137

138

139

TABLE 20.24

140

141

142

143

144

145

146

147

TABLE 20.25

148

149

150

151

152

153

154

155

156

157

158

159

TABLE 20.26

160

161

162

163

164

165

166

167

168

169

170

171

TABLE 20.27

172

173

174

175

176

177

178

179

TABLE 20.28

180

181

182

183

184

185

TABLE 20.29

186

187

188

189

190

191

192

193

TABLE 20.30

194

195

196

197

198

199

TABLE 20.31

200

201

202

203

204

205

206

207

TABLE 20.32

208

209

210

211

212

213

TABLE 20.33

214

215

216

217

218

219

220

221

TABLE 20.34

222

223

224

225

226

227

TABLE 20.35

228

229

230

231

232

233

234

235

TABLE 20.36

236

237

238

239

240

241

242

243

TABLE 20.37

244

245

246

247

TABLE 30.1

67

68

69

70

71

258

248

249

Embodiments of R^(x) include esters, carbamates, carbonates, thioesters,amides, thioamides, and urea groups:

Any reference to the compounds of the invention described heerein alsoincludes a reference to a physiologically acceptable salt thereof.Examples of physiologically acceptable salts of the compounds of theinvention include salts derived from an appropriate base, such as analkali metal or an alkaline earth (for example, Na⁺, Li⁺, K^(+,) Ca⁺²and Mg⁺²), ammonium and NR₄ ⁺ (wherein R is defined herein).Physiologically acceptable salts of a nitrogen atom or an amino groupinclude (a) acid addition salts formed with inorganic acids, forexample, hydrochloric acid, hydrobromic acid, sulfuric acid, sulfamicacids, phosphoric acid, nitric acid and the like; (b) salts formed withorganic acids such as, for example, acetic acid, oxalic acid, tartaricacid, succinic acid, maleic acid, fumaric acid, gluconic acid, citricacid, malic acid, ascorbic acid, benzoic acid, isethionic acid,lactobionic acid, tannic acid, palmitic acid, alginic acid, polyglutamicacid, naphthalenesulfonic acid, methanesulfonic acid, p-toluenesulfonicacid, benzenesulfonic acid, naphthalenedisulfonic acid, polygalacturonicacid, malonic acid, sulfosalicylic acid, glycolic acid,2-hydroxy-3-naphthoate, pamoate, salicylic acid, stearic acid, phthalicacid, mandelic acid, lactic acid, ethanesulfonic acid, lysine, arginine,glutamic acid, glycine, serine, threonine, alanine, isoleucine, leucineand the like; and (c) salts formed from elemental anions for example,chlorine, bromine, and iodine. Physiologically acceptable salts of acompound of a hydroxy group include the anion of said compound incombination with a suitable cation such as Na⁺ and NR₄ ⁺.

For therapeutic use, salts of active ingredients of the compounds of theinvention will be physiologically acceptable, i.e. they will be saltsderived from a physiologically acceptable acid or base. However, saltsof acids or bases which are not physiologically acceptable may also finduse, for example, in the preparation or purification of aphysiologically acceptable compound. All salts, whether or not derivedform a physiologically acceptable acid or base, are within the scope ofthe present invention.

Finally, it is to be understood that the compositions herein comprisecompounds of the invention in their un-ionized, as well as zwitterionicform, and combinations with stoichiometric amounts of water as inhydrates.

The compounds of the invention, exemplified by Formula I-IV may havechiral centers, e.g. chiral carbon or phosphorus atoms. The compounds ofthe invention thus include racemic mixtures of all stereoisomers,including enantiomers, diastereomers, and atropisomers. In addition, thecompounds of the invention include enriched or resolved optical isomersat any or all asymmetric, chiral atoms. In other words, the chiralcenters apparent from the depictions are provided as the chiral isomersor racemic mixtures. Both racemic and diastereomeric mixtures, as wellas the individual optical isomers isolated or synthesized, substantiallyfree of their enantiomeric or diastereomeric partners, are all withinthe scope of the invention. The racemic mixtures are separated intotheir individual, substantially optically pure isomers throughwell-known techniques such as, for example, the separation ofdiastereomeric salts formed with optically active adjuncts, e.g., acidsor bases followed by conversion back to the optically active substances.In most instances, the desired optical isomer is synthesized by means ofstereospecific reactions, beginning with the appropriate stereoisomer ofthe desired starting material.

The term “chiral” refers to molecules which have the property ofnon-superimposability of the mirror image partner, while the term“achiral” refers to molecules which are superimposable on their mirrorimage partner.

The term “stereoisomers” refers to compounds which have identicalchemical constitution, but differ with regard to the arrangement of theatoms or groups in space.

“Diastereomer” refers to a stereoisomer with two or more centers ofchirality and whose molecules are not mirror images of one another.Diastereomers have different physical properties, e.g. melting points,boiling points, spectral properties, reactivities and biologicalproperties. For example, the compounds of Formula I-IV may have a chiralphosphorus atom when R⁷ is

and W¹ and W² are different. When at least one of either W¹ or W² alsohas a chiral center, for example with W¹ or W² is a nitrogen-linked,chiral, naturally occurring α-amino acid ester, then the compound ofFormula I-IV will exists as diastereomers because there are two centersof chirality in the molecule. All such diastereomers and their usesdescribed herein are encompassed by the instant invention. Mixtures ofdiastereomers may be separate under high resolution analyticalprocedures such as electrophoresis, crystallization and/orchromatography. Diastereomeres may have different physical attributessuch as, but not limited to, solubility, chemical stabilities andcrystallinity and may also have different biological properties such as,but not limited to, enzymatic stability, absorption and metabolicstability.

“Enantiomers” refer to two stereoisomers of a compound which arenon-superimposable mirror images of one another.

Stereochemical definitions and conventions used herein generally followS. P. Parker, Ed., McGraw-Hill Dictionary of Chemical Terms (1984)McGraw-Hill Book Company, New York; and Eliel, E. and Wilen, S.,Stereochemistry of Organic Compounds (1994) John Wiley & Sons, Inc., NewYork. Many organic compounds exist in optically active forms, i.e., theyhave the ability to rotate the plane of plane-polarized light. Indescribing an optically active compound, the prefixes D and L or R and Sare used to denote the absolute configuration of the molecule about itschiral center(s). The prefixes d and l, D and L, or (+) and (−) areemployed to designate the sign of rotation of plane-polarized light bythe compound, with S, (−), or l meaning that the compound islevorotatory while a compound prefixed with R, (+), or d isdextrorotatory. For a given chemical structure, these stereoisomers areidentical except that they are mirror images of one another. A specificstereoisomer may also be referred to as an enantiomer, and a mixture ofsuch isomers is often called an enantiomeric mixture. A 50:50 mixture ofenantiomers is referred to as a racemic mixture or a racemate, which mayoccur where there has been no stereoselection or stereospecificity in achemical reaction or process. The terms “racemic mixture” and “racemate”refer to an equimolar mixture of two enantiomeric species, devoid ofoptical activity.

Whenever a compound described herein is substituted with more than oneof the same designated group, e.g., “R” or “R¹”, then it will beunderstood that the groups may be the same or different, i.e., eachgroup is independently selected. Wavy lines,

, indicate the site of covalent bond attachments to the adjoiningsubstructures, groups, moieties, or atoms.

The compounds of the invention can also exist as tautomeric isomers incertain cases. Although only one delocalized resonance structure may bedepicted, all such forms are contemplated within the scope of theinvention. For example, ene-amine tautomers can exist for purine,pyrimidine, imidazole, guanidine, amidine, and tetrazole systems and alltheir possible tautomeric forms are within the scope of the invention.

Methods of Inhibition of a Paramyxoviridae Polymerase

Another aspect of the invention relates to methods of inhibiting theactivity of Paramyxoviridae polymerase comprising the step of treating asample suspected of containing Paramyxoviridae with a composition of theinvention.

Compositions of the invention may act as inhibitors of Paramyxoviridaepolymerase , as intermediates for such inhibitors or have otherutilities as described below. The inhibitors will bind to locations onthe surface or in a cavity of Paramyxoviridae polymerase having ageometry unique to Paramyxoviridae polymerase. Compositions bindingParamyxoviridae polymerase may bind with varying degrees ofreversibility. Those compounds binding substantially irreversibly areideal candidates for use in this method of the invention. Once labeled,the substantially irreversibly binding compositions are useful as probesfor the detection of Paramyxoviridae polymerase. Accordingly, theinvention relates to methods of detecting Paramyxoviridae polymerase ina sample suspected of containing Paramyxoviridae polymerase comprisingthe steps of: treating a sample suspected of containing Paramyxoviridaepolymerase with a composition comprising a compound of the inventionbound to a label; and observing the effect of the sample on the activityof the label. Suitable labels are well known in the diagnostics fieldand include stable free radicals, fluorophores, radioisotopes, enzymes,chemiluminescent groups and chromogens. The compounds herein are labeledin conventional fashion using functional groups such as hydroxyl,carboxyl, sulfhydryl or amino.

Within the context of the invention, samples suspected of containingParamyxoviridae polymerase include natural or man-made materials such asliving organisms; tissue or cell cultures; biological samples such asbiological material samples (blood, serum, urine, cerebrospinal fluid,tears, sputum, saliva, tissue samples, and the like); laboratorysamples; food, water, or air samples; bioproduct samples such asextracts of cells, particularly recombinant cells synthesizing a desiredglycoprotein; and the like. Typically the sample will be suspected ofcontaining an organism which produces Paramyxoviridae polymerase,frequently a pathogenic organism such as a Paramyxoviridae virus.Samples can be contained in any medium including water and organicsolvent\water mixtures. Samples include living organisms such as humans,and man made materials such as cell cultures.

The treating step of the invention comprises adding the composition ofthe invention to the sample or it comprises adding a precursor of thecomposition to the sample. The addition step comprises any method ofadministration as described above.

If desired, the activity of Paramyxoviridae polymerase after applicationof the composition can be observed by any method including direct andindirect methods of detecting Paramyxoviridae polymerase activity.Quantitative, qualitative, and semiquantitative methods of determiningParamyxoviridae polymerase activity are all contemplated. Typically oneof the screening methods described above are applied, however, any othermethod such as observation of the physiological properties of a livingorganism are also applicable.

Organisms that contain Paramyxoviridae polymerase include theParamyxoviridae virus. The compounds of this invention are useful in thetreatment or prophylaxis of Paramyxoviridae infections in animals or inman.

However, in screening compounds capable of inhibiting humanParamyxoviridae viruses, it should be kept in mind that the results ofenzyme assays may not correlate with cell culture assays. Thus, a cellbased assay should be the primary screening tool.

Screens for Paramyxoviridae Polymerase Inhibitors.

Compositions of the invention are screened for inhibitory activityagainst Paramyxoviridae polymerase by any of the conventional techniquesfor evaluating enzyme activity. Within the context of the invention,typically compositions are first screened for inhibition ofParamyxoviridae polymerase in vitro and compositions showing inhibitoryactivity are then screened for activity in vivo. Compositions having invitro Ki (inhibitory constants) of less then about 5×10⁻⁶ M andpreferably less than about 1×10⁻⁷ M are preferred for in vivo use.

Useful in vitro screens have been described in detail and will not beelaborated here. However, the examples describe suitable in vitroassays.

Pharmaceutical Formulations

The compounds of this invention are formulated with conventionalcarriers and excipients, which will be selected in accord with ordinarypractice. Tablets will contain excipients, glidants, fillers, bindersand the like. Aqueous formulations are prepared in sterile form, andwhen intended for delivery by other than oral administration generallywill be isotonic. All formulations will optionally contain excipientssuch as those set forth in the “Handbook of Pharmaceutical Excipients”(1986). Excipients include ascorbic acid and other antioxidants,chelating agents such as EDTA, carbohydrates such as dextran,hydroxyalkylcellulose, hydroxyalkylmethylcellulose, stearic acid and thelike. The pH of the formulations ranges from about 3 to about 11, but isordinarily about 7 to 10.

While it is possible for the active ingredients to be administered aloneit may be preferable to present them as pharmaceutical formulations. Theformulations, both for veterinary and for human use, of the inventioncomprise at least one active ingredient, as above defined, together withone or more acceptable carriers therefor and optionally othertherapeutic ingredients, particularly those additional therapeuticingredients as discussed herein. The carrier(s) must be “acceptable” inthe sense of being compatible with the other ingredients of theformulation and physiologically innocuous to the recipient thereof.

The formulations include those suitable for the foregoing administrationroutes. The formulations may conveniently be presented in unit dosageform and may be prepared by any of the methods well known in the art ofpharmacy. Techniques and formulations generally are found in Remington'sPharmaceutical Sciences (Mack Publishing Co., Easton, Pa.). Such methodsinclude the step of bringing into association the active ingredient withthe carrier which constitutes one or more accessory ingredients. Ingeneral the formulations are prepared by uniformly and intimatelybringing into association the active ingredient with liquid carriers orfinely divided solid carriers or both, and then, if necessary, shapingthe product.

Formulations of the present invention suitable for oral administrationmay be presented as discrete units such as capsules, cachets or tabletseach containing a predetermined amount of the active ingredient; as apowder or granules; as a solution or a suspension in an aqueous ornon-aqueous liquid; or as an oil-in-water liquid emulsion or awater-in-oil liquid emulsion. The active ingredient may also beadministered as a bolus, electuary or paste.

A tablet is made by compression or molding, optionally with one or moreaccessory ingredients. Compressed tablets may be prepared by compressingin a suitable machine the active ingredient in a free-flowing form suchas a powder or granules, optionally mixed with a binder, lubricant,inert diluent, preservative, surface active or dispersing agent. Moldedtablets may be made by molding in a suitable machine a mixture of thepowdered active ingredient moistened with an inert liquid diluent. Thetablets may optionally be coated or scored and optionally are formulatedso as to provide slow or controlled release of the active ingredienttherefrom.

For infections of the eye or other external tissues e.g. mouth and skin,the formulations are preferably applied as a topical ointment or creamcontaining the active ingredient(s) in an amount of, for example, 0.075to 20% w/w (including active ingredient(s) in a range between 0.1% and20% in increments of 0.1% w/w such as 0.6% w/w, 0.7% w/w, etc.),preferably 0.2 to 15% w/w and most preferably 0.5 to 10% w/w. Whenformulated in an ointment, the active ingredients may be employed witheither a paraffinic or a water-miscible ointment base. Alternatively,the active ingredients may be formulated in a cream with an oil-in-watercream base.

If desired, the aqueous phase of the cream base may include, forexample, at least 30% w/w of a polyhydric alcohol, i.e. an alcoholhaving two or more hydroxyl groups such as propylene glycol, butane1,3-diol, mannitol, sorbitol, glycerol and polyethylene glycol(including PEG 400) and mixtures thereof. The topical formulations maydesirably include a compound which enhances absorption or penetration ofthe active ingredient through the skin or other affected areas. Examplesof such dermal penetration enhancers include dimethyl sulphoxide andrelated analogs.

The oily phase of the emulsions of this invention may be constitutedfrom known ingredients in a known manner. While the phase may comprisemerely an emulsifier (otherwise known as an emulgent), it desirablycomprises a mixture of at least one emulsifier with a fat or an oil orwith both a fat and an oil. Preferably, a hydrophilic emulsifier isincluded together with a lipophilic emulsifier which acts as astabilizer. It is also preferred to include both an oil and a fat.Together, the emulsifier(s) with or without stabilizer(s) make up theso-called emulsifying wax, and the wax together with the oil and fatmake up the so-called emulsifying ointment base which forms the oilydispersed phase of the cream formulations.

Emulgents and emulsion stabilizers suitable for use in the formulationof the invention include Tween® 60, Span® 80, cetostearyl alcohol,benzyl alcohol, myristyl alcohol, glyceryl mono-stearate and sodiumlauryl sulfate.

The choice of suitable oils or fats for the formulation is based onachieving the desired cosmetic properties. The cream should preferablybe a non-greasy, non-staining and washable product with suitableconsistency to avoid leakage from tubes or other containers. Straight orbranched chain, mono- or dibasic alkyl esters such as di-isoadipate,isocetyl stearate, propylene glycol diester of coconut fatty acids,isopropyl myristate, decyl oleate, isopropyl palmitate, butyl stearate,2-ethylhexyl palmitate or a blend of branched chain esters known asCrodamol CAP may be used, the last three being preferred esters. Thesemay be used alone or in combination depending on the propertiesrequired. Alternatively, high melting point lipids such as white softparaffin and/or liquid paraffin or other mineral oils are used.

Pharmaceutical formulations according to the present invention comprisea combination according to the invention together with one or morepharmaceutically acceptable carriers or excipients and optionally othertherapeutic agents. Pharmaceutical formulations containing the activeingredient may be in any form suitable for the intended method ofadministration. When used for oral use for example, tablets, troches,lozenges, aqueous or oil suspensions, dispersible powders or granules,emulsions, hard or soft capsules, syrups or elixirs may be prepared.Compositions intended for oral use may be prepared according to anymethod known to the art for the manufacture of pharmaceuticalcompositions and such compositions may contain one or more agentsincluding sweetening agents, flavoring agents, coloring agents andpreserving agents, in order to provide a palatable preparation. Tabletscontaining the active ingredient in admixture with non-toxicpharmaceutically acceptable excipient which are suitable for manufactureof tablets are acceptable. These excipients may be, for example, inertdiluents, such as calcium or sodium carbonate, lactose, calcium orsodium phosphate; granulating and disintegrating agents, such as maizestarch, or alginic acid; binding agents, such as starch, gelatin oracacia; and lubricating agents, such as magnesium stearate, stearic acidor talc. Tablets may be uncoated or may be coated by known techniquesincluding microencapsulation to delay disintegration and adsorption inthe gastrointestinal tract and thereby provide a sustained action over alonger period. For example, a time delay material such as glycerylmonostearate or glyceryl distearate alone or with a wax may be employed.

Formulations for oral use may be also presented as hard gelatin capsuleswhere the active ingredient is mixed with an inert solid diluent, forexample calcium phosphate or kaolin, or as soft gelatin capsules whereinthe active ingredient is mixed with water or an oil medium, such aspeanut oil, liquid paraffin or olive oil.

Aqueous suspensions of the invention contain the active materials inadmixture with excipients suitable for the manufacture of aqueoussuspensions. Such excipients include a suspending agent, such as sodiumcarboxymethylcellulose, methylcellulose, hydroxypropyl methylcelluose,sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia,and dispersing or wetting agents such as a naturally-occurringphosphatide (e.g., lecithin), a condensation product of an alkyleneoxide with a fatty acid (e.g., polyoxyethylene stearate), a condensationproduct of ethylene oxide with a long chain aliphatic alcohol (e.g.,heptadecaethyleneoxycetanol), a condensation product of ethylene oxidewith a partial ester derived from a fatty acid and a hexitol anhydride(e.g., polyoxyethylene sorbitan monooleate). The aqueous suspension mayalso contain one or more preservatives such as ethyl or n-propylp-hydroxy-benzoate, one or more coloring agents, one or more flavoringagents and one or more sweetening agents, such as sucrose or saccharin.

Oil suspensions may be formulated by suspending the active ingredient ina vegetable oil, such as arachis oil, olive oil, sesame oil or coconutoil, or in a mineral oil such as liquid paraffin. The oral suspensionsmay contain a thickening agent, such as beeswax, hard paraffin or cetylalcohol. Sweetening agents, such as those set forth above, and flavoringagents may be added to provide a palatable oral preparation. Thesecompositions may be preserved by the addition of an antioxidant such asascorbic acid.

Dispersible powders and granules of the invention suitable forpreparation of an aqueous suspension by the addition of water providethe active ingredient in admixture with a dispersing or wetting agent, asuspending agent, and one or more preservatives. Suitable dispersing orwetting agents and suspending agents are exemplified by those disclosedabove. Additional excipients, for example sweetening, flavoring andcoloring agents, may also be present.

The pharmaceutical compositions of the invention may also be in the formof oil-in-water emulsions. The oily phase may be a vegetable oil, suchas olive oil or arachis oil, a mineral oil, such as liquid paraffin, ora mixture of these. Suitable emulsifying agents includenaturally-occurring gums, such as gum acacia and gum tragacanth,naturally-occurring phosphatides, such as soybean lecithin, esters orpartial esters derived from fatty acids and hexitol anhydrides, such assorbitan monooleate, and condensation products of these partial esterswith ethylene oxide, such as polyoxyethylene sorbitan monooleate. Theemulsion may also contain sweetening and flavoring agents. Syrups andelixirs may be formulated with sweetening agents, such as glycerol,sorbitol or sucrose. Such formulations may also contain a demulcent, apreservative, a flavoring or a coloring agent.

The pharmaceutical compositions of the invention may be in the form of asterile injectable preparation, such as a sterile injectable aqueous oroleaginous suspension. This suspension may be formulated according tothe known art using those suitable dispersing or wetting agents andsuspending agents which have been mentioned above. The sterileinjectable preparation may also be a sterile injectable solution orsuspension in a non-toxic parenterally acceptable diluent or solvent,such as a solution in 1,3-butane-diol or prepared as a lyophilizedpowder. Among the acceptable vehicles and solvents that may be employedare water, Ringer's solution and isotonic sodium chloride solution. Inaddition, sterile fixed oils may conventionally be employed as a solventor suspending medium. For this purpose any bland fixed oil may beemployed including synthetic mono- or diglycerides. In addition, fattyacids such as oleic acid may likewise be used in the preparation ofinjectables.

The amount of active ingredient that may be combined with the carriermaterial to produce a single dosage form will vary depending upon thehost treated and the particular mode of administration. For example, atime-release formulation intended for oral administration to humans maycontain approximately 1 to 1000 mg of active material compounded with anappropriate and convenient amount of carrier material which may varyfrom about 5 to about 95% of the total compositions (weight:weight). Thepharmaceutical composition can be prepared to provide easily measurableamounts for administration. For example, an aqueous solution intendedfor intravenous infusion may contain from about 3 to 500 μg of theactive ingredient per milliliter of solution in order that infusion of asuitable volume at a rate of about 30 mL/hr can occur.

Formulations suitable for topical administration to the eye also includeeye drops wherein the active ingredient is dissolved or suspended in asuitable carrier, especially an aqueous solvent for the activeingredient. The active ingredient is preferably present in suchformulations in a concentration of 0.5 to 20%, advantageously 0.5 to10%, and particularly about 1.5% w/w.

Formulations suitable for topical administration in the mouth includelozenges comprising the active ingredient in a flavored basis, usuallysucrose and acacia or tragacanth; pastilles comprising the activeingredient in an inert basis such as gelatin and glycerin, or sucroseand acacia; and mouthwashes comprising the active ingredient in asuitable liquid carrier.

Formulations for rectal administration may be presented as a suppositorywith a suitable base comprising for example cocoa butter or asalicylate.

Formulations suitable for intrapulmonary or nasal administration have aparticle size for example in the range of 0.1 to 500 microns, such as0.5, 1, 30, 35 etc., which is administered by rapid inhalation throughthe nasal passage or by inhalation through the mouth so as to reach thealveolar sacs. Suitable formulations include aqueous or oily solutionsof the active ingredient. Formulations suitable for aerosol or drypowder administration may be prepared according to conventional methodsand may be delivered with other therapeutic agents such as compoundsheretofore used in the treatment or prophylaxis of Paramyxoviridaeinfections as described below.

In another aspect, the invention is a novel, efficacious, safe,nonirritating and physiologically compatible inhalable compositioncomprising a compound of Formula I-IV, or a pharmaceutically acceptablesalt thereof, suitable for treating Paramyxoviridae infections andpotentially associated bronchiolitis. Preferred pharmaceuticallyacceptable salts are inorganic acid salts including hydrochloride,hydrobromide, sulfate or phosphate salts as they may cause lesspulmonary irritation. Preferably, the inhalable formulation is deliveredto the endobronchial space in an aerosol comprising particles with amass median aerodynamic diameter (MMAD) between about 1 and about 5 μm.Preferably, the compound of Formula I-IV is formulated for aerosoldelivery using a nebulizer, pressurized metered dose inhaler (pMDI), ordry powder inhaler (DPI).

Non-limiting examples of nebulizers include atomizing, jet, ultrasonic,pressurized, vibrating porous plate, or equivalent nebulizers includingthose nebulizers utilizing adaptive aerosol delivery technology (Denyer,J. Aerosol medicine Pulmonary Drug Delivery 2010, 23 Supp 1, S1-S10). Ajet nebulizer utilizes air pressure to break a liquid solution intoaerosol droplets. An ultrasonic nebulizer works by a piezoelectriccrystal that shears a liquid into small aerosol droplets. A pressurizednebulization system forces solution under pressure through small poresto generate aerosol droplets. A vibrating porous plate device utilizesrapid vibration to shear a stream of liquid into appropriate dropletsizes.

In a preferred embodiment, the formulation for nebulization is deliveredto the endobronchial space in an aerosol comprising particles with aMMAD predominantly between about 1 μm and about 5 μm using a nebulizerable to aerosolize the formulation of the compound of Formula I-IV intoparticles of the required MMAD. To be optimally therapeuticallyeffective and to avoid upper respiratory and systemic side effects, themajority of aerosolized particles should not have a MMAD greater thanabout 5 μm. If an aerosol contains a large number of particles with aMMAD larger than 5 μm, the particles are deposited in the upper airwaysdecreasing the amount of drug delivered to the site of inflammation andbronchoconstriction in the lower respiratory tract. If the MMAD of theaerosol is smaller than about 1 μm, then the particles have a tendencyto remain suspended in the inhaled air and are subsequently exhaledduring expiration.

When formulated and delivered according to the method of the invention,the aerosol formulation for nebulization delivers a therapeuticallyefficacious dose of the compound of Formula I-IV to the site ofParamyxoviridae infection sufficient to treat the Paramyxoviridaeinfection. The amount of drug administered must be adjusted to reflectthe efficiency of the delivery of a therapeutically efficacious dose ofthe compound of Formula I-IV. In a preferred embodiment, a combinationof the aqueous aerosol formulation with the atomizing, jet, pressurized,vibrating porous plate, or ultrasonic nebulizer permits, depending onthe nebulizer, about, at least, 20, to about 90%, typically about 70%delivery of the administered dose of the compound of Formula I-IV intothe airways. In a preferred embodiment, at least about 30 to about 50%of the active compound is delivered. More preferably, about 70 to about90% of the active compound is delivered.

In another embodiment of the instant invention, a compound of FormulaI-IV or a pharmaceutically acceptable salt thereof, is delivered as adry inhalable powder. The compounds of the invention are administeredendobronchially as a dry powder formulation to efficacious deliver fineparticles of compound into the endobronchial space using dry powder ormetered dose inhalers. For delivery by DPI, the compound of Formula I-IVis processed into particles with, predominantly, MMAD between about 1 μmand about 5 μm by milling spray drying, critical fluid processing, orprecipitation from solution. Media milling, jet milling and spray-dryingdevices and procedures capable of producing the particle sizes with aMMAD between about 1 μm and about 5 μm are well know in the art. In oneembodiment, excipients are added to the compound of Formula I-IV beforeprocessing into particles of the required sizes. In another embodiment,excipients are blended with the particles of the required size to aid indispersion of the drug particles, for example by using lactose as anexcipient.

Particle size determinations are made using devices well known in theart. For example a multi-stage Anderson cascade impactor or othersuitable method such as those specifically cited within the USPharmacopoeia Chapter 601 as characterizing devices for aerosols withinmetered-dose and dry powder inhalers.

In another preferred embodiment, a compound of Formula I-IV is deliveredas a dry powder using a device such as a dry powder inhaler or other drypowder dispersion devices. Non-limiting examples of dry powder inhalersand devices include those disclosed in U.S. Pat. Nos. 5,458,135;5,740,794; 5,775,320; 5,785,049; 3,906,950; 4,013,075; 4,069,819;4,995,385; 5,522,385; 4,668,218; 4,667,668; 4,805,811 and 5,388,572.There are two major designs of dry powder inhalers. One design is ametering device in which a reservoir for the drug is place within thedevice and the patient adds a dose of the drug into the inhalationchamber. The second design is a factory-metered device in which eachindividual dose has been manufactured in a separate container. Bothsystems depend on the formulation of the drug into small particles ofMMAD from 1 μm and about 5 μm, and often involve co-formulation withlarger excipient particles such as, but not limited to, lactose. Drugpowder is placed in the inhalation chamber (either by device metering orby breakage of a factory-metered dosage) and the inspiratory flow of thepatient accelerates the powder out of the device and into the oralcavity. Non-laminar flow characteristics of the powder path cause theexcipient-drug aggregates to decompose, and the mass of the largeexcipient particles causes their impaction at the back of the throat,while the smaller drug particles are deposited deep in the lungs. Inpreferred embodiments, a compound of Formula I-IV, or a pharmaceuticallyacceptable salt thereof, is delivered as a dry powder using either typeof dry powder inhaler as described herein, wherein the MMAD of the drypowder, exclusive of any excipients, is predominantly in the range of 1μm to about 5 μm.

In another preferred embodiment, a compound of Formula I-IV is deliveredas a dry powder using a metered dose inhaler. Non-limiting examples ofmetered dose inhalers and devices include those disclosed in U.S. Pat.Nos. 5,261,538; 5,544,647; 5,622,163; 4,955,371; 3,565,070; 3,361,306and 6,116,234. In preferred embodiments, a compound of Formula I-IV, ora pharmaceutically acceptable salt thereof, is delivered as a dry powderusing a metered dose inhaler wherein the MMAD of the dry powder,exclusive of any excipients, is predominantly in the range of about 1-5μm.

Formulations suitable for vaginal administration may be presented aspessaries, tampons, creams, gels, pastes, foams or spray formulationscontaining in addition to the active ingredient such carriers as areknown in the art to be appropriate.

Formulations suitable for parenteral administration include aqueous andnon-aqueous sterile injection solutions which may contain anti-oxidants,buffers, bacteriostats and solutes which render the formulation isotonicwith the blood of the intended recipient; and aqueous and non-aqueoussterile suspensions which may include suspending agents and thickeningagents.

The formulations are presented in unit-dose or multi-dose containers,for example sealed ampoules and vials, and may be stored in afreeze-dried (lyophilized) condition requiring only the addition of thesterile liquid carrier, for example water for injection, immediatelyprior to use. Extemporaneous injection solutions and suspensions areprepared from sterile powders, granules and tablets of the kindpreviously described. Preferred unit dosage formulations are thosecontaining a daily dose or unit daily sub-dose, as herein above recited,or an appropriate fraction thereof, of the active ingredient.

It should be understood that in addition to the ingredients particularlymentioned above the formulations of this invention may include otheragents conventional in the art having regard to the type of formulationin question, for example those suitable for oral administration mayinclude flavoring agents.

The invention further provides veterinary compositions comprising atleast one active ingredient as above defined together with a veterinarycarrier therefor.

Veterinary carriers are materials useful for the purpose ofadministering the composition and may be solid, liquid or gaseousmaterials which are otherwise inert or acceptable in the veterinary artand are compatible with the active ingredient. These veterinarycompositions may be administered orally, parenterally or by any otherdesired route.

Compounds of the invention are used to provide controlled releasepharmaceutical formulations containing as active ingredient one or morecompounds of the invention (“controlled release formulations”) in whichthe release of the active ingredient are controlled and regulated toallow less frequency dosing or to improve the pharmacokinetic ortoxicity profile of a given active ingredient.

Effective dose of active ingredient depends at least on the nature ofthe condition being treated, toxicity, whether the compound is beingused prophylactically (lower doses) or against an active viralinfection, the method of delivery, and the pharmaceutical formulation,and will be determined by the clinician using conventional doseescalation studies. It can be expected to be from about 0.0001 to about100 mg/kg body weight per day; typically, from about 0.01 to about 10mg/kg body weight per day; more typically, from about 0.01 to about 5mg/kg body weight per day; most typically, from about 0.05 to about 0.5mg/kg body weight per day. For example, the daily candidate dose for anadult human of approximately 70 kg body weight will range from 1 mg to1000 mg, preferably between 5 mg and 500 mg, and may take the form ofsingle or multiple doses.

Routes of Administration

One or more compounds of the invention (herein referred to as the activeingredients) are administered by any route appropriate to the conditionto be treated. Suitable routes include oral, rectal, nasal, pulmonary,topical (including buccal and sublingual), vaginal and parenteral(including subcutaneous, intramuscular, intravenous, intradermal,intrathecal and epidural), and the like. It will be appreciated that thepreferred route may vary with for example the condition of therecipient. An advantage of the compounds of this invention is that theyare orally bioavailable and can be dosed orally.

Combination Therapy

Compositions of the invention are also used in combination with otheractive ingredients. For the treatment of Paramyxoviridae virusinfections, preferably, the other active therapeutic agent is activeagainst Paramyxoviridae virus infections, particularly respiratorysyncytial virus infections and/or parainfluenza virus infections.Non-limiting examples of these other active therapeutic agents areribavirin, palivizumab, motavizumab, RSV-IGIV (RespiGam®), MEDI-557,A-60444, MDT-637, BMS-433771, and mixtures thereof.

Many of the infections of the Paramyxoviridae viruses are respiratoryinfections. Therefore, additional active therapeutics used to treatrespiratory symptoms and sequelae of infection may be used incombination with the compounds of Formula I-IV. The additional agentsare preferably administered orally or by direct inhalation. For example,other preferred additional therapeutic agents in combination with thecompounds of Formula I-IV for the treatment of viral respiratoryinfections include, but are not limited to, bronchodilators andcorticosteroids.

Glucocorticoids, which were first introduced as an asthma therapy in1950 (Carryer, Journal of Allergy, 21, 282-287, 1950), remain the mostpotent and consistently effective therapy for this disease, althoughtheir mechanism of action is not yet fully understood (Morris, J.Allergy Clin. Immunol., 75 (1 Pt) 1-13, 1985). Unfortunately, oralglucocorticoid therapies are associated with profound undesirable sideeffects such as truncal obesity, hypertension, glaucoma, glucoseintolerance, acceleration of cataract formation, bone mineral loss, andpsychological effects, all of which limit their use as long-termtherapeutic agents (Goodman and Gilman, 10th edition, 2001). A solutionto systemic side effects is to deliver steroid drugs directly to thesite of inflammation. Inhaled corticosteroids (ICS) have been developedto mitigate the severe adverse effects of oral steroids. Non-limitingexamples of corticosteroids that may be used in combinations with thecompounds of Formula I-IV are dexamethasone, dexamethasone sodiumphosphate, fluorometholone, fluorometholone acetate, loteprednol,loteprednol etabonate, hydrocortisone, prednisolone, fludrocortisones,triamcinolone, triamcinolone acetonide, betamethasone, beclomethasonediproprionate, methylprednisolone, fluocinolone, fluocinolone acetonide,flunisolide, fluocortin-21-butylate, flumethasone, flumetasone pivalate,budesonide, halobetasol propionate, mometasone furoate, fluticasonepropionate, ciclesonide; or a pharmaceutically acceptable salts thereof.

Other anti-inflammatory agents working through anti-inflammatory cascademechanisms are also useful as additional therapeutic agents incombination with the compounds of Formula I-IV for the treatment ofviral respiratory infections. Applying “anti-inflammatory signaltransduction modulators” (referred to in this text as AISTM), likephosphodiesterase inhibitors (e.g. PDE-4, PDE-5, or PDE-7 specific),transcription factor inhibitors (e.g. blocking NFKB through IKKinhibition), or kinase inhibitors (e.g. blocking P38 MAP, JNK, PI3K,EGFR or Syk) is a logical approach to switching off inflammation asthese small molecules target a limited number of common intracellularpathways—those signal transduction pathways that are critical points forthe anti-inflammatory therapeutic intervention (see review by P. J.Barnes, 2006). These non-limiting additional therapeutic agents include:5-(2,4-Difluoro-phenoxy)-1-isobutyl-1H-indazole-6-carboxylic acid(2-dimethylamino-ethyl)-amide (P38 Map kinase inhibitor ARRY-797);3-Cyclopropylmethoxy-N-(3,5-dichloro-pyridin-4-yl)-4-difluorormethoxy-benzamide(PDE-4 inhibitor Roflumilast);4-[2-(3-cyclopentyloxy-4-methoxyphenyl)-2-phenyl-ethyl]-pyridine (PDE-4inhibitor CDP-840);N-(3,5-dichloro-4-pyridinyl)-4-(difluoromethoxy)-8-[(methylsulfonyl)amino]-1-dibenzofurancarboxamide(PDE-4 inhibitor Oglemilast);N-(3,5-Dichloro-pyridin-4-yl)-2-[1-(4-fluorobenzyl)-5-hydroxy-1H-indol-3-yl]-2-oxo-acetamide(PDE-4 inhibitor AWD 12-281);8-Methoxy-2-trifluoromethyl-quinoline-5-carboxylic acid(3,5-dichloro-1-oxy-pyridin-4-yl)-amide (PDE-4 inhibitor Sch 351591);4-[5-(4-Fluorophenyl)-2-(4-methanesulfinyl-phenyl)-1H-imidazol-4-yl]-pyridine(P38 inhibitor SB-203850);4-[4-(4-Fluoro-phenyl)-1-(3-phenyl-propyl)-5-pyridin-4-yl-1H-imidazol-2-yl]-but-3-yn-1-ol(P38 inhibitor RWJ-67657);4-Cyano-4-(3-cyclopentyloxy-4-methoxy-phenyl)-cyclohexanecarboxylic acid2-diethyl amino-ethyl ester (2-diethyl-ethyl ester prodrug ofCilomilast, PDE-4 inhibitor);(3-Chloro-4-fluorophenyl)-[7-methoxy-6-(3-morpholin-4-yl-propoxy)-quinazolin-4-yl]-amine(Gefitinib, EGFR inhibitor); and4-(4-Methyl-piperazin-1-ylmethyl)-N-[4-methyl-3-(4-pyridin-3-yl-pyrimidin-2-ylamino)-phenyl]-benzamide(Imatinib, EGFR inhibitor).

Combinations comprising inhaled β2-adrenoreceptor agonistbronchodilators such as formoterol, albuterol or salmeterol with thecompounds of Formula I-IV are also suitable, but non-limiting,combinations useful for the treatment of respiratory viral infections.

Combinations of inhaled β2-adrenoreceptor agonist bronchodilators suchas formoterol or salmeterol with ICS's are also used to treat both thebronchoconstriction and the inflammation (Symbicort® and Advair®,respectively). The combinations comprising these ICS andβ2-adrenoreceptor agonist combinations along with the compounds ofFormula I-IV are also suitable, but non-limiting, combinations usefulfor the treatment of respiratory viral infections.

For the treatment or prophylaxis of pulmonary broncho-constriction,anticholinergics are of potential use and, therefore, useful as anadditional therapeutic agents in combination with the compounds ofFormula I-IV for the treatment of viral respiratory infections. Theseanticholinergics include, but are not limited to, antagonists of themuscarinic receptor (particularly of the M3 subtype) which have showntherapeutic efficacy in man for the control of cholinergic tone in COPD(Witek, 1999);1-{4-Hydroxy-1-[3,3,3-tris-(4-fluoro-phenyl)-propionyl]-pyrrolidine-2-carbonyl}-pyrrolidine-2-carboxylicacid (1-methyl-piperidin-4-ylmethyl)-amide;3-[3-(2-Diethylamino-acetoxy)-2-phenyl-propionyloxy]-8-isopropyl-8-methyl-8-azonia-bicyclo[3.2.1]octane(Ipratropium-N,N-diethylglycinate);1-Cyclohexyl-3,4-dihydro-1H-isoquinoline-2-carboxylic acid1-aza-bicyclo[2.2.2]oct-3-yl ester (Solifenacin);2-Hydroxymethyl-4-methanesulfinyl-2-phenyl-butyric acid1-aza-bicyclo[2.2.2]oct-3-yl ester (Revatropate);2-{1-[2-(2,3-Dihydro-benzofuran-5-yl)-ethyl]-pyrrolidin-3-yl}-2,2-diphenyl-acetamide(Darifenacin); 4-Azepan-1-yl-2,2-diphenyl-butyramide (Buzepide);7-[3-(2-Diethylamino-acetoxy)-2-phenyl-propionyloxy]-9-ethyl-9-methyl-3-oxa-9-azonia-tricyclo[3.3.1.02,4]nonane(Oxitropium-N,N-diethylglycinate);7-[2-(2-Diethylamino-acetoxy)-2,2-di-thiophen-2-yl-acetoxy]-9,9-dimethyl-3-oxa-9-azonia-tricyclo[3.3.1.02,4]nonane(Tiotropium-N,N-diethylglycinate); Dimethylamino-acetic acid2-(3-diisopropylamino-1-phenyl-propyl)-4-methyl-phenyl ester(Tolterodine-N,N-dimethylglycinate);3-[4,4-Bis-(4-fluoro-phenyl)-2-oxo-imidazolidin-1-yl]-1-methyl-1-(2-oxo-2-pyridin-2-yl-ethyl)-pyrrolidinium;1-[1-(3-Fluoro-benzyl)-piperidin-4-yl]-4,4-bis-(4-fluoro-phenyl)-imidazolidin-2-one;1-Cyclooctyl-3-(3-methoxy-1-aza-bicyclo[2.2.2]oct-3-yl)-1-phenyl-prop-2-yn-1-ol;3-[2-(2-Diethylamino-acetoxy)-2,2-di-thiophen-2-yl-acetoxy]-1-(3-phenoxy-propyl)-1-azonia-bicyclo[2.2.2]octane(Aclidinium-N,N-diethylglycinate); or(2-Diethylamino-acetoxy)-di-thiophen-2-yl-acetic acid1-methyl-1-(2-phenoxy-ethyl)-piperidin-4-yl ester.

The compounds of Formula I-IV may also be combined with mucolytic agentsto treat both the infection and symptoms of respiratory infections. Anon-limiting example of a mucolytic agent is ambroxol. Similarly, thecompounds of Formula I-IV may be combined with expectorants to treatboth the infection and symptoms of respiratory infections. Anon-limiting example of an expectorant is guaifenesin.

Nebulized hypertonic saline is used to improve immediate and long-termclearance of small airways in patients with lung diseases (Kuzik, J.Pediatrics 2007, 266). The compounds of Formula I-IV may also becombined with nebulized hypertonic saline particularly when theParamyxoviridae virus infection is complicated with bronchiolitis. Thecombination of the compounds of Formula I-IV with hypertonic saline mayalso comprise any of the additional agents discussed above. In apreferred aspect, nebulized about 3% hypertonic saline is used.

It is also possible to combine any compound of the invention with one ormore additional active therapeutic agents in a unitary dosage form forsimultaneous or sequential administration to a patient. The combinationtherapy may be administered as a simultaneous or sequential regimen.When administered sequentially, the combination may be administered intwo or more administrations.

Co-administration of a compound of the invention with one or more otheractive therapeutic agents generally refers to simultaneous or sequentialadministration of a compound of the invention and one or more otheractive therapeutic agents, such that therapeutically effective amountsof the compound of the invention and one or more other activetherapeutic agents are both present in the body of the patient.

Co-administration includes administration of unit dosages of thecompounds of the invention before or after administration of unitdosages of one or more other active therapeutic agents, for example,administration of the compounds of the invention within seconds,minutes, or hours of the administration of one or more other activetherapeutic agents. For example, a unit dose of a compound of theinvention can be administered first, followed within seconds or minutesby administration of a unit dose of one or more other active therapeuticagents. Alternatively, a unit dose of one or more other therapeuticagents can be administered first, followed by administration of a unitdose of a compound of the invention within seconds or minutes. In somecases, it may be desirable to administer a unit dose of a compound ofthe invention first, followed, after a period of hours (e.g., 1-12hours), by administration of a unit dose of one or more other activetherapeutic agents. In other cases, it may be desirable to administer aunit dose of one or more other active therapeutic agents first,followed, after a period of hours (e.g., 1-12 hours), by administrationof a unit dose of a compound of the invention.

The combination therapy may provide “synergy” and “synergistic”, i.e.the effect achieved when the active ingredients used together is greaterthan the sum of the effects that results from using the compoundsseparately. A synergistic effect may be attained when the activeingredients are: (1) co-formulated and administered or deliveredsimultaneously in a combined formulation; (2) delivered by alternationor in parallel as separate formulations; or (3) by some other regimen.When delivered in alternation therapy, a synergistic effect may beattained when the compounds are administered or delivered sequentially,e.g. in separate tablets, pills or capsules, or by different injectionsin separate syringes. In general, during alternation therapy, aneffective dosage of each active ingredient is administered sequentially,i.e. serially, whereas in combination therapy, effective dosages of twoor more active ingredients are administered together. A synergisticanti-viral effect denotes an antiviral effect which is greater than thepredicted purely additive effects of the individual compounds of thecombination.

In still yet another embodiment, the present application provides formethods of inhibiting Paramyxoviridae polymerase in a cell, comprising:contacting a cell infected with HCV with an effective amount of acompound of Formula I-IV, or a pharmaceutically acceptable salt,solvate, and/or ester thereof, whereby Paramyxoviridae polymerase isinhibited.

In still yet another embodiment, the present application provides formethods of inhibiting Paramyxoviridae polymerase in a cell, comprising:contacting a cell infected with HCV with an effective amount of acompound of Formula I-IV, or a pharmaceutically acceptable salt,solvate, and/or ester thereof, and at least one additional activetherapeutic agent, whereby Paramyxoviridae polymerase is inhibited.

In still yet another embodiment, the present application provides formethods of inhibiting Paramyxoviridae polymerase in a cell, comprising:contacting a cell infected with Paramyxoviridae virus with an effectiveamount of a compound of Formula I-IV, or a pharmaceutically acceptablesalt, solvate, and/or ester thereof, and at least one additional activetherapeutic agent selected

In still yet another embodiment, the present application provides formethods of treating Paramyxoviridae virus infection in a patient,comprising: administering to the patient a therapeutically effectiveamount of a compound of Formula I-IV, or a pharmaceutically acceptablesalt, solvate, and/or ester thereof.

In still yet another embodiment, the present application provides formethods of treating Paramyxoviridae virus infection in a patient,comprising: administering to the patient a therapeutically effectiveamount of a compound of Formula I-IV, or a pharmaceutically acceptablesalt, solvate, and/or ester thereof, and at least one additional activetherapeutic agent, whereby Paramyxoviridae polymerase is inhibited.

In still yet another embodiment, the present application provides formethods of treating Paramyxoviridae virus infection in a patient,comprising: administering to the patient a therapeutically effectiveamount of a compound of Formula I-IV, or a pharmaceutically acceptablesalt, solvate, and/or ester thereof, and at least one additional activetherapeutic agent.

Metabolites of the Compounds of the Invention

Also falling within the scope of this invention are the in vivometabolic products of the compounds described herein, to the extent suchproducts are novel and unobvious over the prior art. Such products mayresult for example from the oxidation, reduction, hydrolysis, amidation,esterification and the like of the administered compound, primarily dueto enzymatic processes. Accordingly, the invention includes novel andunobvious compounds produced by a process comprising contacting acompound of this invention with a mammal for a period of time sufficientto yield a metabolic product thereof. Such products typically areidentified by preparing a radiolabelled (e.g. ¹⁴C or ³H) compound of theinvention, administering it parenterally in a detectable dose (e.g.greater than about 0.5 mg/kg) to an animal such as rat, mouse, guineapig, monkey, or to man, allowing sufficient time for metabolism to occur(typically about 30 seconds to 30 hours) and isolating its conversionproducts from the urine, blood or other biological samples. Theseproducts are easily isolated since they are labeled (others are isolatedby the use of antibodies capable of binding epitopes surviving in themetabolite). The metabolite structures are determined in conventionalfashion, e.g. by MS or NMR analysis. In general, analysis of metabolitesis done in the same way as conventional drug metabolism studieswell-known to those skilled in the art. The conversion products, so longas they are not otherwise found in vivo, are useful in diagnostic assaysfor therapeutic dosing of the compounds of the invention even if theypossess no HCV polymerase inhibitory activity of their own.

Recipes and methods for determining stability of compounds in surrogategastrointestinal secretions are known. Compounds are defined herein asstable in the gastrointestinal tract where less than about 50 molepercent of the protected groups are deprotected in surrogate intestinalor gastric juice upon incubation for 1 hour at 37° C. Simply because thecompounds are stable to the gastrointestinal tract does not mean thatthey cannot be hydrolyzed in vivo. The prodrugs of the inventiontypically will be stable in the digestive system but may besubstantially hydrolyzed to the parental drug in the digestive lumen,liver or other metabolic organ, or within cells in general.

EXAMPLES

Certain abbreviations and acronyms are used in describing theexperimental details. Although most of these would be understood by oneskilled in the art, Table 1 contains a list of many of theseabbreviations and acronyms.

TABLE 1 List of abbreviations and acronyms. Abbreviation Meaning Ac₂Oacetic anhydride AIBN 2,2’-azobis(2-methylpropionitrile) Bn benzyl BnBrbenzylbromide BSA bis(trimethylsilyl)acetamide BzCl benzoyl chloride CDIcarbonyl diimidazole DABCO 1,4-diazabicyclo[2.2.2]octane DBN1,5-diazabicyclo[4.3.0]non-5-ene DDQ2,3-dichloro-5,6-dicyano-1,4-benzoquinone DBU1,5-diazabicyclo[5.4.0]undec-5-ene DCA dichloroacetamide DCCdicyclohexylcarbodiimide DCM dichloromethane DMAP4-dimethylaminopyridine DME 1,2-dimethoxyethane DMTCl dimethoxytritylchloride DMSO dimethylsulfoxide DMTr 4, 4’-dimethoxytrityl DMFdimethylformamide EtOAc ethyl acetate ESI electrospray ionization HMDShexamethyldisilazane HPLC High pressure liquid chromatography LDAlithium diisopropylamide LRMS low resolution mass spectrum MCPBAmeta-chloroperbenzoic acid MeCN acetonitrile MeOH methanol MMTC monomethoxytrityl chloride m/z or m/e mass to charge ratio MH⁺ mass plus 1MH⁻ mass minus 1 MsOH methanesulfonic acid MS or ms mass spectrum NBSN-bromosuccinimide Ph phenyl rt or r.t. room temperature TBAFtetrabutylammonium fluoride TMSC1 chlorotrimethylsilane TMSBrbromotrimethylsilane TMSI iodotrimethylsilane TMSOTf(trimethylsilyl)trifluoromethylsulfonate TEA triethylamine TBAtributylamine TBAP tributylammonium pyrophosphate TBSClt-butyldimethylsilyl chloride TEAB triethylammonium bicarbonate TFAtrifluoroacetic acid TLC or tlc thin layer chromatography Trtriphenylmethyl Tol 4-methylbenzoyl Turbo Grignard 1:1 mixture ofisopropylmagnesium chloride and lithium chloride δ parts per milliondown field from tetramethylsilane

Preparation of Compounds (2S)-ethyl2-(chloro(phenoxy)phosphorylamino)propanoate (Chloridate A)

Ethyl alanine ester hydrochloride salt (1.69 g, 11 mmol) was dissolvedin anhydrous CH₂Cl₂ (10 mL) and the mixture stirred with cooling to 0°C. under N₂(g). Phenyl dichlorophosphate (1.49 mL, 10 mmol) was addedfollowed by dropwise addition of Et₃N over 10 min. The reaction mixturewas then slowly warmed to RT and stirred for 12 h. Anhydrous Et₂O (50mL) was added and the mixture stirred for 30 min. The solid that formedwas removed by filtration, and the filtrate concentrated under reducedpressure. The residue was subjected to silica gel chromatography elutingwith 0-50% EtOAc in hexanes to provide intermediate A (1.13 g, 39%).

¹H NMR (300 MHz, CDCl₃) δ 7.39-7.27 (m, 5H), 4.27 (m, 3H), 1.52 (m, 3H),1.32 (m, 3H).

³¹P NMR (121.4 MHz, CDCl₃) δ 8.2, 7.8.

(2S)-2-ethylbutyl 2-(chloro(phenoxy)phosphorylamino)propanoate(Chloridate B)

The 2-ethylbutyl alanine chlorophosphoramidate ester B was preparedusing the same procedure as chloridate A except substituting2-ethylbutyl alanine ester for ethyl alanine ester. The material is usedcrude in the next reaction. Treatment with methanol or ethanol forms thedisplaced product with the requisite LCMS signal.

(2S)-isopropyl 2-(chloro(phenoxy)phosphorylamino)propanoate (ChloridateC)

The isopropyl alanine chlorophosphoramidate ester C was prepared usingthe same procedure as chloridate A except substituting isopropyl alanineester for the ethyl alanine ester. The material is used crude in thenext reaction. Treatment with methanol or ethanol forms the displacedproduct with the requisite LCMS signal.

(2R,3R,4S,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-3,4-dihydroxy-5-(hydroxymethyl)tetrahydrofuran-2-carbonitrile(Compound 1)

The commercially available lactol (10 g, 23.8 mmol) was dissolved inanhydrous DMSO (30 mL) under N₂(g). Ac₂O (20 mL) was added and theresultant reaction mixture stirred at RT for 48 h. The reaction mixturewas poured onto ice H₂O (500 mL) and the mixture stirred for 20 min. Themixture was extracted with EtOAc (3×200 mL) and the combined organicextracts were then washed with H₂O (3×200 mL). The organic extract wasdried over anhydrous MgSO₄, filtered and concentrated under reducedpressure. The residue was dissolved in CH₂Cl₂ and subjected to silicagel chromatography eluting with 25% EtOAc in hexanes to provide thelactone (9.55 g, 96%).

¹H NMR (400 MHz, DMSO) δ□□ 7.30-7.34 (m, 13H), 7.19-7.21 (m, 2H),4.55-4.72 (m, 6H), 4.47 (s, 2H), 4.28 (d, J=3.9 Hz,1H), 3.66 (m, 2H).

LCMS m/z 436.1 [M+H₂O], 435.2 [M+OH]− Tr=2.82 min

HPLC Tr=4.59 [2-98% ACN in H2) over 5 min @2 ml/min flow.

The bromopyrazole (prepared according to WO2009/132135) (0.5 g, 2.4mmol) was suspended in anhydrous THF (10 mL) under N₂(g). The suspensionwas stirred and TMSCl (0.67 mL, 5.28 mmol) was added. The mixture wasstirred for 20 min. at RT and then cooled to −78° C. after which time asolution of n-BuLi (6 mL, 1.6 N in hexanes, 9.6 mmol) was added slowly.The reaction mixture was stirred for 10 min. at −78° C. and then thelactone (1 g, 2.4 mmol) was added via syringe. When the reaction wascomplete as measured by LCMS, AcOH was added to quench the reaction. Themixture was concentrated under reduced pressure and the residuedissolved in a mixture of CH₂Cl₂ and H₂O (100 mL, 1:1). The organiclayer was separated and washed with H₂O (50 mL). The organic layer wasthen dried over anhydrous MgSO₄, filtered and concentrated under reducedpressure. The residue was subjected to silica gel chromatography elutingwith 0-50% EtOAc in hexanes to provide the product as a 1:1 mixture ofanomers (345 mg, 26% yield). LCMS m/z 553 [M+H].

The hydroxy nucleoside (1.1 g, 2.0 mmol) was dissolved in anhydrousCH₂Cl₂ (40 mL) and the solution cooled with stirring to 0° C. underN₂(g). TMSCN (0.931 mL, 7 mmol) was added and the mixture stirred for afurther 10 min. TMSOTf (1.63 mL, 9.0 mmol) was slowly added to thereaction and the mixture stirred for 1 h. The reaction mixture was thendiluted with CH₂Cl₂ (120 mL) and aqueous NaHCO₃ (120 mL) was added toquench the reaction. The reaction mixture was stirred for a further 10min and the organic layer separated. The aqueous layer was extractedwith CH₂Cl₂ (150 mL) and the combined organic extracts dried overanhydrous MgSO₄, filtered and concentrated under reduced pressure. Theresidue was dissolved in a minimal amount of CH₂Cl₂ and subjected tosilica gel chromatography eluting with a gradient of 0-75% EtOAc andhexanes to provide the tribenzyl cyano nucleoside as a mixture ofanomers. (0.9 g, 80%).

¹H NMR (300 MHz, CD₃CN) δ 7.94 (s, 0.5H), 7.88 (s, 0.5H), 7.29-7.43 (m,13H), 7.11-7.19 (m, 1H), 6.82-6.88 (m,1H), 6.70-6.76 (m, 1H), 6.41 (bs,2H), 5.10 (d, J=3.9 Hz, 0.5H), 4.96 (d, J=5.1 Hz, 0.5H), 4.31-4.85 (m,7H), 4.09-4.18 (m, 2H), 3.61-3.90 (m, 2H).

LCMS m/z 562 [M+H].

The tribenzyl cyano nucleoside (70 mg, 0.124 mmol) was dissolved inanhydrous CH₂Cl₂ (2 mL) and cooled to −78° C. under N₂(g). A solution ofBCl₃ (1N in CH₂Cl₂, 0.506 mL, 0.506 mmol) was added and the reactionmixture stirred for 1 h. at −78° C. When the reaction was complete byLC/MS, MeOH was added to quench the reaction. The reaction mixture wasallowed to warm to room RT and the solvent removed under reducedpressure. The residue was subjected to C18 reverse phase HPLC, elutingfor 5 min with H₂O (0.1% TFA), followed by a gradient of 0-70% MeCN inH₂O (0.1% TFA) over 35 min, to elute the α-anomer (20 mg, 37%), andβ-anomer 1 (20 mg, 37%).

(α-anomer)

¹H NMR (300 MHz, D₂O) δ 7.96 (s, 1H), 7.20 (d, J=4.8 Hz, 1H), 6.91 (d,J=4.8 Hz, 1H), 4.97 (d, J=4.4 Hz, 1H), 4.56-4.62 (m, 1H), 4.08-4.14 (m,1H), 3.90 (dd, J=12.9, 2.4 Hz, 1H), 3.70 (dd, J=13.2, 4.5 Hz, 1H).

(β-anomer)

¹H NMR (400 MHz, DMSO) δ 7.91 (s, 1H), 7.80-8.00 (br s, 2H), 6.85-6.89(m, 2H), 6.07 (d, J=6.0 Hz, 1H), 5.17 (br s, 1H), 4.90 (br s, 1H), 4.63(t, J=3.9 Hz, 1H), 4.02-4.06 (m, 1H), 3.94 (br s, 1H), 3.48-3.64 (m,2H).

LCMS m/z 292.2 [M+H], 290.0 [M−H]. Tr=0.35 min.

13C NMR (400 MHZ, DMSO), 156.0, 148.3, 124.3, 117.8, 117.0, 111.2,101.3, 85.8, 79.0, 74.7, 70.5, 61.4

HPLC Tr=1.32 min

(2R,3R,4R,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-3-fluoro-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-carbonitrile(Compound 2)

2-Deoxy-2-fluoro-4,5-O,O-dibenzyl-D-arabinose

1′-Methoxy-2-deoxy-2-fluoro-4,5-O,O-dibenzyl-D-arabinose (1.0 g, 2.88mmol) in TFA (13.5 mL) was treated with H₂O (1.5 mL) and the resultantmixture stirred for 5 h. The mixture was then diluted with EtOAc (100mL) and treated with saturated NaHCO₃ (50 mL). The organic layer wasseparated and washed with NaCl (50 mL), dried over anhydrous MgSO₄,filtered and concentrated under reduced pressure. The residue wassubjected to silica gel chromatography (80 g SiO₂ Combiflash HP GoldColumn) eluting with 0-100% EtOAc in hexanes to afford2-deoxy-2-fluoro-4,5-O,O-dibenzyl-D-arabinose (695 mg, 72%) as a whitesolid: R_(f)=0.52 (25% EtOAc in hexanes);

¹H NMR (300 MHz, CDCl₃) δ 7.30 (m, 10H), 5.35 (m, 1H), 4.68-4.29 (m,7H), 3.70 (d, J=10.5 Hz, 1H), 3.50 (d, J=10.5 Hz, 2H).

¹⁹F NMR (282.2 MHz, CDCl₃) δ −207 (m), −211 (m).

LCMS m/z 350 [M+H₂O].

(3R,4R,5R)-4-(benzyloxy)-5-(benzyloxymethyl)-3-fluorodihydrofuran-2(311)-one

2-Deoxy-2-fluoro-4, 5-O,O-dibenzyl-D-arabinose (4.3 g, 12.8 mmol) wasdissolved in CH₂Cl₂ (85 mL) was treated with 4 Å MS (10 g) andpyridinium dichromate (14.4 g, 38.3 mmol). The resultant mixture wasstirred for 24 h and then filtered through a pad of Celite. The eluantwas concentrated under reduced pressure and the residue subjected tosilica gel chromatography (120 g SiO₂ HP Gold Combiflash Column) elutingwith 0-100% EtOAc in hexanes to afford(3R,4R,5R)-4-(benzyloxy)-5-(benzyloxymethyl)-3-fluorodihydrofuran-2(3H)-oneas a clear oil (3.5 g, 83%): R_(f)=0.25 (25% EtOAc in hexanes).

¹H NMR (300 MHz, CDCl₃) δ 7.37 (m, 10H), 5.45 (dd, J=49, 5.7, Hz, 1H),4.85 (d, J=11.7 Hz, 1H), 4.52 (m, 4 H), 4.29 (d, J=5.4 Hz, 1H), 2.08(dd, J=15.3, 10.2 Hz, 2H).

¹⁹F NMR (282.2 MHz, CDCl₃) δ −216.

LCMS m/z 348 [M+H₂O].

HPLC (6-98% MeCN-H₂O gradient, 0.05% TFA modifier) t_(R)=5.29 min.

Phenomenex Synergi 4 m Hydro-RP 80 A, 50×4.60 mm, 4 micron; 2 mL/minflow rate

(3R,4R,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-(benzyloxy)-5-(benzyloxymethyl)-3-fluorotetrahydrofuran-2-ol

7-Bromopyrrolo[1,2-f][1,2,4]-triazin-4-amine (68 mg, 0.319 mmol) in THF(1.4 mL) was treated with TMSCl (89 μL, 0.703 mmol) and the mixturestirred for 2 h. The mixture was then cooled to −78° C. and treated withnBuLi (1.0 M in hexanes, 1.09 mL, 1.09 mmol). The solution was stirredfor 30 min and then treated with(3R,4R,5R)-4-(benzyloxy)-5-(benzyloxymethyl)-3-fluorodihydrofuran-2(3H)-one(106 mg, 0.319 mmol) dropwise in THF (1.4 mL). The resultant mixture wasstirred for 30 min and then AcOH (83 μL, 1.44 mmol) in THF (1.0 mL) wasadded to quench the reaction. The mixture was warmed to RT and thenconcentrated under reduced pressure. The residue was diluted with EtOAc(100 mL) and washed with saturated NaCl solution (50 mL). The organiclayer was dried over anhydrous MgSO₄, filtered and concentrated underreduced pressure. The residue was subjected to silica gel chromatography(40 g SiO₂ HP Gold Combiflash Column) eluting with 0-100% EtOAc inhexanes followed by a 0-100% gradient of (20% MeOH in EtOAc) in EtOAc toafford(3R,4R,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-(benzyloxy)-5-(benzyloxymethyl)-3-fluorotetrahydrofuran-2-olas a white solid (68 mg, 44%, 60/40 mixture of α/β isomers). R_(f)=0.32(EtOAc).

¹H NMR (300 MHz, CDCl₃) δ 8.05 (s, 1H), 7.86 (s, 1H), 7.81 (s, 1H), 7.64(s, 1H), 7.26 (m, 10H), 6.95 (m, 1H), 6.71 (m, 1H), 6.08 (m, 1H), 5.34(m, 1H), 4.65 (m, 6H), 4.71 (m, 2H).

¹⁹F NMR (282.2 MHz, CDCl₃) δ −211 (m).

LCMS m/z 465 [M+H].

HPLC (6-98% MeCN—H₂O gradient, 0.05% TFA modifier) t_(R)=4.37 min.(α-isomer), 4.54 min. (β-isomer).

(3R,4R,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-(benzyloxy)-5-(benzyloxymethyl)-3-fluorotetrahydrofuran-2-carbonitrile

(3R,4R,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-(benzyloxy)-5-(benzyloxymethyl)-3-fluorotetrahydrofuran-2-ol(195 mg, 0.42 mmol) was dissolved in MeCN (1.4 mL) was treated withTMSCN (336 μL, 2.52 mmol) and In(OTf)₃ (708 mg, 1.26 mmol). The solutionwas stirred at 70° C. for 18 h and then cooled to 0° C. The mixture wastreated with saturated NaHCO₃ solution (20 drops) then warmed to RT anddiluted with EtOAc (100 mL) and H₂O (50 mL). The organic layer wasseparated and washed with saturated NaCl solution (50 mL), dried overMgSO₄, filtered and concentrated under reduced pressure. The residue wassubjected to silica gel chromatography (40 g SiO₂ HP Gold CombiflashColumn) eluting with 0-100% EtOAc in hexanes to afford(3R,4R,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-(benzyloxy)-5-(benzyloxymethyl)-3-fluorotetrahydrofuran-2-carbonitrileas a white solid (110 mg, 55%, 60/40 mixture of α/β isomers). Data forboth isomers: R_(f)=0.53 (EtOAc).

¹H NMR (300 MHz, CDCl₃) δ 8.01 (s, 1H), 7.94 (s, 1H), 7.30 (m, 10H),7.00 (d, J=4.5 Hz, 1H), 6.93 (d, J=4.8 Hz, 1H), 6.87 (d, J=5.4 Hz, 1H),6.70 (d, J=4.8 Hz, 1H), 5.85 (dd, J=52, 3.3 Hz, 1H), 5.55 (dd, J=53, 4.5Hz, 1H), 4.71 (m, 7H), 3.87 (m, 2H), 3.72 (m, 2H).

¹⁹F NMR (282.2 MHz, CDCl₃) δ −196 (m), 203 (m).

LCMS m/z 474 [M+H].

HPLC (6-98% MeCN—H₂O gradient, 0.05% TFA modifier) t_(R)=4.98 min.

(2R,3R,4R,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-3-fluoro-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-carbonitrile(2)

(3R,4R,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-(benzyloxy)-5-(benzyloxymethyl)-3-fluorotetrahydrofuran-2-carbonitrile(110 mg, 0.23 mmol) was dissolved in CH₂Cl₂ (1.5 mL) and cooled to 0° C.The reaction mixture was treated with BCl₃ (1.0 M in CH₂Cl₂, 766 μL,0.77 mmol) and stirred for 2 h. The mixture was then cooled to −78° C.and treated with Et₃N (340 μL, 2.44 mmol) followed by MeOH (2 mL) beforeallowing to warm to RT. The reaction was concentrated under reducedpressure and then co-evaporated with MeOH (3×5 mL). The residue was thensuspended in H₂O (5 mL) and treated with NaHCO₃ (1 g). The solution wasstirred for 10 min and then concentrated under reduced pressure. Theresidue was filtered and washed with MeOH (3×10 mL) on a fritted glassfunnel (coarse) and the eluant concentrated under reduced pressure. Theresidue was subjected to reverse phase HPLC (6-98% MeCN in H₂O gradientwith 0.05% TFA modifier) to afford(2R,3R,4R,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-3-fluoro-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-carbonitrile2 as a white solid (16.8 mg, 25%) and the α-isomer.

Data for the β-isomer: R_(f)=0.13 (10% MeOH in EtOAc).

¹H NMR (300 MHz, CD₃OD) δ 8.09 (s, 1H), 7.28 (d, J=5.1 Hz, 1H), 7.17 (d,J=5.1 Hz, 1H), 5.42 (dd, J=53, 3.3 Hz, 1H), 4.20 (m, 2H), 3.99 (d, J=3.6Hz, 1H), 3.77 (d, J=3.6 Hz, 1H).

¹⁹F NMR (282.2 MHz, CDCl₃) δ −197 (m).

LCMS m/z 294 [M+H].

HPLC (2-98% MeCN—H₂O gradient, 0.05% TFA modifier) t_(R)=1.49 min.

(2R,3R,4R,5S)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-fluoro-2-(hydroxymethyl)-5-methyltetrahydrofuran-3-ol(Compound 3)

The starting nucleoside (prepared as described in the synthesis ofcompound 2) (0.355 g, 0.765 mmol) was dissolved in anhydrous THF (35 mL)and cooled to 0° C. with stirring under N₂(g). A solution of methylmagnesium chloride (2 mL, 6 mmol) (3N in THF) was added and theresultant mixture stirred overnight. Acetic acid (7 mmol) was added toquench the reaction and then the solvents were removed by rotory underreduced pressure. The residue was re-dissolved in CH₂Cl₂ and thesolution subjected to a plug of silica gel to isolate the product (0.355g) as a crude mixture. LC/MS (m/z: 480, M⁺¹). The crude material wasdissolved in anhydrous CH₂Cl₂ (20 mL) and placed under N₂(g). Thesolution was stirred and treated with methanesulfonic acid (0.2 mL, 2.74mmol). The reaction mixture was stirred for 12 h at RT and then quenchedby the addition of Et₃N (3.5 mmol). The mixture was concentrated underreduced pressure and the residue subjected to silica gel chromatographyto provide the methyl substituted nucleoside (0.174 g, 0.377 mmol, 44%yield) as a 4:1 mixture of beta- and alpha-anomers respectively.

¹H NMR (300 MHz, CD₃CN) major anomer δ 7.87 (s, 1H), 7.27-7.40 (m, 10H), 6.77 (d, J=4.5 HZ, 1H), 6.70 (d, J=4.5 Hz, 1H), 6.23 (br s, 2H),5.53 (dd, J=55, 3.3 Hz, 1H), 4.42-4.75 (m, 4H), 4.19-4.26 (m, 1H),3.65-4.00 (m, 3H), 1.74 (d, J=3.9 Hz, 3H).

¹⁹F NMR (282.2 MHz, CD₃CN) major anomer δ −207 (m, 1F)

LCMS m/z 463 [M+H].

The benzylated nucleoside material (0.134 g, 0.290 mmol), Degussacatalyst (0.268 g) and AcOH (30 mL) were mixed together. The reactionatmosphere was charged with H₂ (g) and the reaction stirred for 2 h. Thecatalyst was removed by filtration and the mixture concentrated underreduced pressure. The residue was dissolved in a minimal amount of H₂Oand subjected to reverse phase HPLC (C¹⁸ hydro RP column) to isolate theβ-anomer 3 (0.086 g, 0.217 mmol, 57% yield).

¹H NMR (300 MHz, D₂O) δ□7.87 (s, 1H), 7.22 (d, J=4.8 Hz, 1H), 6.87 (d,J=4.8 Hz, 1H), 5.35 (dd, J=54, 3.6 Hz, 1H), 3.97-4.10 (m, 2H), 3.81 (dd,J=12.6, 2.1 Hz, 1H), 3.64 (dd, J=12.6, 4.8 Hz, 1H), 1.65 (d, J=4.2 Hz,3H).

¹⁹F NMR (282.2 MHz, CD₃CN) δ□ −207 (m, 1F).

A small amount of alpha anomer was characterized as follows.

¹H NMR (300 MHz, D₂O) δ□ 7.86 (s, 1H), 7.26 (d, J=4.8 Hz, 1H), 6.85 (d,J=4.8 Hz, 1H), 5.31 (dd, J=54, 3.9 Hz, 1H), 4.39 (ddd, J=26.1, 9.9, 3.6Hz, 2H), 4.00-4.05 (m, 1H), 3.90 (dd, J=12.3, 2.1 Hz, 1H), 3.66 (dd,J=12.6, 4.8, 1H), 1.56 (s, 3H).

¹⁹F NMR (282.2 MHz, CD₃CN) δ□ −198 (dd, J=54, 26 Hz, 1F).

(2R)-isopropyl2-((((2R,3R,4R,5S)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-fluoro-3-hydroxy-5-methyltetrahydrofuran-2-yl)methoxy)-(phenoxy)phosphorylamino)propanoate(Compound 4)

The nucleoside 3 (0.011 g, 0.04 mmol) was dissolved intrimethylphosphate (2 mL) and cooled to 0° C. The mixture was stirredunder an atmosphere of N₂(g) and 1-Methylimidazole (0.320 mL, 5 mmol)followed by the alaninylmonoisopropyl, monophenol phosphorchloridate C(0.240 mL, 4.4 mmol) was added. The reaction mixture was stirred for 2h. at 0° C. and then allowed to warm slowly to RT. while monitoring byLC/MS. When complete by LCMS, the reaction mixture was treated with H₂O(5 mL) and then concentrated under reduced pressure. The residue wasdissolved in CH₂Cl₂ and subjected to silica gel chromatography elutingwith 0-100% EtOAc in hexanes. The product fractions were collected andconcentrated. The residue was subjected to prep HPLC to yield thealanine isopropyl monoamidate prodrug 4 as a mixture of isomers (4.7 mg,0.003 mmol, 6%).

¹H NMR (300 MHz, CD3CN) δ 7.87 (s, 1H), 7.17-7.44 (m, 5 H), 6.71-6.83(m, 2H), 6.14 (br, s, 2H), 5.38 (dd, J=56, 3.3 Hz, 1H), 4.92-5.01 (m,1H), 3.86-4.46 (m, 6H), 3.58 (m, 1H), 1.73 (m, 3H), 1.18-1.34 (m, 9H)

LCMS m/z 552 [M+H].

(2R)-ethyl2-((((2R,3R,4R,5S)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-fluoro-3-hydroxy-5-methyltetrahydrofuran-2-yl)methoxy)(phenoxy)phosphorylamino)propanoate(Compound 5)

The nucleoside 3 (0.026 g, 0.092 mmol) was dissolved intrimethylphosphate (2 mL) and cooled to 0° C. The mixture was stirredunder N₂(g) and 1-methylimidazole (0.062 mL, 0.763 mmol) followed by thechloridate A (0.160 g, 0.552 mmol) were added. The reaction mixture wasstirred for 2 h. at 0° C. and then allowed to warm slowly to RT. H₂O (5mL) was added to quench the reaction and then the mixture concentratedunder reduced pressure. The residue was dissolved in CH₂Cl₂ andsubjected to silica gel chromatography eluting with 0-100% EtOAc inhexanes. The product fractions were collected and concentrated. Crudeproduct was eluted using 0 to 100 percent EtOAc in hexanes. The crudeproduct was collected and concentrated under reduced pressure. Theresidue was subjected to prep HPLC to yield 5 (2.0 mg, 4% yield).

LCMS m/z 538 [M+H].

((2R,3R,4R,5S)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-4-fluoro-3-hydroxy-5-methyltetrahydrofuran-2-yl)methyltetrahydrogen triphosphate (Compound 6)

The nucleoside 3 (0.022 g, 0.056 mmol) was dissolved intrimethylphosphate (1 mL) and stirred under N₂(g). Phosphorousoxychloride (0.067 mL, 0.73 mmol) was added and the mixture stirred for2 h. Monitoring by analytical ion-exchange column determined the time atwhich >80 percent of monophosphate was formed. A solution oftributylamine (0.44 mL, 1.85 mmol) and triethylammonium pyrophosphate(0.327 g, 0.72 mmol) dissolved in anhydrous DMF (1 mL) was added. Thereaction mixture was stirred for 20 min and then quenched by theaddition of 1N triethylammonium bicarbonate solution in H₂O (5 mL). Themixture was concentrated under reduced pressure and the residuere-dissolved in H₂O. The solution was subjected to ion exchangechromatography to yield the title product 6 (1.7 mg, 6% yield).

LCMS m/z 521 [M−H]. Tr=0.41

HPLC ion exchange TR=9.40 min

(2R,3R,5S)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-3-hydroxy-5-(hydroxymethyl)-tetrahydrofuran-2-carbonitrile(Compound 7)

((3αR,5S,6αR)-2,2-dimethyl-tetrahydrofuro[2,3-d][1,3]dioxol-5-yl)methanol

The acetate material (1.2 g, 5.5 mmol) (J. Org. Chem. 1985, 50, 3547, DeBernardo et al) was dissolved in a 1:1 mixture MeOH and THF (10 mL). A1N solution of NaOH(aq) (10 mL) was added until the pH was 13. Thereaction mixture was stirred for 2 h and then neutralized to pH 8-9 bythe addition of AcOH. The mixture was extracted with EtOAc (10×30 mL)and the combined organic extracts dried over anhydrous Na₂SO₄, filteredand concentrated under reduced pressure. The residue was subjected tosilica gel chromatography eluting with 0-70% EtOAc in hexanes to givethe desired product (866 mg, 90%).

¹H NMR (300 MHz, CDCl₃) δ 5.84 (d, J=3.6 Hz, 1H), 4.78 (t, J=4.5 Hz,1H), 4.38 (m, 1H), 3.93-3.54 (m, 2H), 2.04-1.84 (m, 2H), 1.52 (s, 3H),1.33 (s, 3H).

(3αR,5S,6αR)-5-(benzyloxymethyl)-2,2-dimethyl-tetrahydrofuro[2,3-d][1,3]dioxole

Sodium hydride (188 mg, 7.46 mmol) was dissolved in anhydrous THF (5 mL)and stirred under N₂(g) at RT. The alcohol (866 mg, 4.97 mmol) wasdissolved in anhydrous THF (3 mL) and then added in portions over 5 min.to the sodium hydride mixture. The resultant mixture was stirred for 20min. and then benzyl bromide (892 μL, 7.46 mmol) was added. The reactionwas stirred for 2 h and then poured onto a mixture of ice cold aqueousNaHCO₃ and EtOAc (30 mL). The organic layer was separated and then theaqueous layer re-extracted with EtOAc (30 mL). The combined organicextracts were dried over anhydrous Na₂SO₄, filtered and concentratedunder reduced pressure. The residue was subjected to silica gelchromatography eluting with 0-40% EtOAc in hexanes to give the benzylether product (912 mg, 69%).

¹H NMR (300 MHz, CDCl₃) δ 7.35-7.27 (m, 5H), 5.86 (d, J=3.6 Hz, 1H),4.74 (t, J=4.2 Hz, 1H), 4.60 (s, 2H), 4.42 (m, 1H), 3.69-3.53 (m, 2H),2.10-2.04 (m, 1H), 1.83-1.77 (m, 1H), 1.52 (s, 3H), 1.33 (s, 3H).

(3R,5S)-5-(benzyloxymethyl)-tetrahydrofuran-2,3-diol

The benzyl ether (910 mg, 3.44 mmol) was dissolved in a 1:1 AcOH and H₂O(20 mL) mixture and stirred at 60° C. for 7 h. The mixture wasconcentrated under reduced pressure and the residue subjected to silicagel chromatography eluting with 0-70% EtOAc in hexanes to give the diolproduct (705 mg, 91%).

¹H NMR (300 MHz, CDCl₃) δ 7.36-7.27 (m, 5H), 5.40 (d, J=3.9 Hz, 0.5H),5.17 (s, 0.5H), 4.67-4.56 (m, 3H), 4.33 (m, 0.5H), 4.24 (d, J=4.8 Hz,0.5H), 3.71-3.67 (m, 1H), 3.56-3.42 (m, 2H), 2.31-2.22 (m, 1H),2.08-1.89 (m, 2H).

(3R,5S)-5-(benzyloxymethyl)-3-hydroxy-dihydrofuran-2(3H)-one

The diol (705 mg, 3.14 mmol) was dissolved in benzene (30 mL) andtreated with a silver carbonate celite mixture (3.46 g, 6.28 mmol). Theresultant mixture was stirred at 80° C. under N₂(g) for 2 h. The mixturewas then cooled to RT, filtered and concentrated under reduced pressure.The residue was subjected to silica gel chromatography eluting with0-70% EtOAc in hexanes to give the lactone product (600 mg, 86%).

¹H NMR (300 MHz, CDCl₃) δ 7.39-7.27 (m, 5H), 4.75-4.68 (m, 1H),4.60-4.49 (m, 2H), 3.74-3.54 (m, 2H), 2.61-2.35 (m, 2H), 2.38-2.28 (m,1H).

(3R, 5S)-3-(benzyloxy)-5-(benzyloxymethyl)-dihydrofuran-2(3H)-one

The lactone (600 mg, 2.7 mmol) was dissolved in EtOAc (30 mL) andtreated with silver oxide (626 mg, 2.7 mmol) followed by benzyl bromide(387 μL, 3.24 mmol). The reaction mixture was then stirred at 50° C.under N₂(g) for 8 h. Additional silver oxide (300 mg) was then added andthe resultant mixture stirred at 50° C. for 16 h. Additional benzylbromide (50 uL) and silver oxide (150 mg) were added and the mixturestirred for an additional 8 h. The reaction mixture was allowed to cool,filtered and then concentrated under reduced pressure. The residue wassubjected to silica gel chromatography eluting with 0-20% EtOAc inhexanes to give the title product (742 mg, 88%).

¹H NMR (300 MHz, CDCl₃) δ 7.39-7.27 (m, 10H), 4.99 (d, J=11.4 Hz, 1H),4.72 (m, 2H), 4.56 (m, 2H), 4.39 (t, J=8.1 Hz, 1H), 3.72-3.51 (m, 2H),2.42-2.25 (m, 2H).

(3R,5S)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-3-(benzyloxy)-5-(benzyloxymethyl)-tetrahydrofuran-2-ol

The 7-bromopyrrolo[1,2-f][1,2,4]triazin-4-amine (607 mg, 2.85 mmol) wasdissolved in anhydrous THF (10 mL) and stirred under Ar(g) at RT. TMSCl(1.1 mL, 8.55 mmol) was added dropwise and the mixture stirred for 2 h.The reaction was concentrated under reduced pressure and then driedunder high vacuum. The residue was suspended in THF (20 mL) and stirredunder Ar(g) at −78° C. A 2.5M n-BuLi solution in hexane (2.28 mL, 5.7mmol) was added dropwise over 10 min. and the resultant mixture stirredfor 60 min. The lactone (742 mg, 2.37 mmol) dissolved in anhydrous THF(7 mL) was added to the above mixture over 20 min. The reaction mixturewas stirred for 2 h. and then quenched with AcOH until pH was 5-6. Themixture was allowed to warm to RT and then diluted with EtOAc. Thesolution was washed with saturated NaHCO₃ solution, saturated NaCl,dried over anhydrous Na₂SO₄ and concentrated under reduced pressure. Theresidue was subjected to silica gel chromatography eluting with 0-80%EtOAc in hexanes to give the title product (250 mg, 24%).

LCMS m/z 447.2 [M+H], 445.1 [M−H].

(3R,5S)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-3-(benzyloxy)-5-(benzyloxymethyl)-tetrahydrofuran-2-carbonitrile

The alcohol (250 mg, 0.56 mmol) was dissolved in anhydrous CH₂Cl₂(10 mL)and stirred under Ar(g) at −15° C. TMSCN (448 μL, 3.36 mmol) was addeddropwise and the mixture stirred for 10 min. TMSOTf (466 μL, 2.58 mmol)was added dropwise over 10 min and the resultant mixture stirred for 90min. at −15° C. Additional TMSCN (224 μL, 3 eq.) and TMSOTf (202 μL, 2eq.) was added and stirring continued for 5 h. Saturated aqueous NaHCO₃solution was added to quench the reaction and the mixture stirred for 10min. The organic layer was separated and washed with saturated aqueousNaHCO₃ solution, saturated NaCl solution, dried over anhydrous Na₂SO₄,filtered and concentrated under reduced pressure. The residue wassubjected to silica gel chromatography eluting with 0-70% EtOAc inhexanes to give the title product (150 mg, 59%).

LCMS m/z 456.3 [M+H], 454.1 [M−H].

(2R,3R,5S)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-3-hydroxy-5-(hydroxymethyl)-tetrahydrofuran-2-carbonitrile(7)

The benzyl ether (150 mg, 0.329 mmol) was dissolved in anhydrous CH₂Cl₂(2 mL) and the mixture stirred under Ar(g) at −20° C. A 1M BCl₃ solutionin CH₂Cl₂ (724 μL, 0.724 mmol) was added dropwise and the resultantmixture stirred for 2 h. Additional 1M BCl₃ in CH₂Cl₂ (724 μL, 0.724mmol) was added and stirring continued for 2 h. The mixture was thencooled to −78° C. and slowly treated with a 2:1 mixture of Et₃N and MeOH(3 mL). The mixture was stirred for 10 min and then treated with MeOH(10 mL). The reaction was allowed to warm to RT and then concentratedunder reduced pressure. The residue was dissolved in MeOH andconcentrated under reduced pressure. The residue was dissolved in MeOHagain and treated with solid NaHCO₃. The mixture was stirred for 5 minand then the solid removed by filtration. The solution was concentratedunder reduced pressure and subjected to preparative HPLC to provide thedesired product 7 (10 mg, 11%).

¹H NMR (300 MHz, D₂O) δ 7.71 (s, 1H), 6.75 (d, J=4.5 Hz, 1H), 6.65 (d,J=4.8 Hz, 1H), 4.91 (t, J=6.3 Hz, 1H), 4.57 (m, 1H), 3.67-3.47 (m, 2H),2.18 (m, 2H).

LCMS m/z 276.1 [M+H], 274.0 [M−H].

(2S)-isopropyl2-((((2R,3S,4R,5R)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-cyano-3,4-dihydroxytetrahydrofuran-2-yl)methoxy)(phenoxy)-phosphorylamino)propanoate(Compound 8)

The nucleoside 1 (45mg, 0.15mmol) was dissolved in anhydrous trimethylphosphate (0.5 mL) and the solution stirred under N₂(g) at 0° C. Methylimidazole (36 μL, 0.45 mmol) was added to the solution.Chlorophosphoramidate C (69 mg, 0.225 mmol) was dissolved in anhydrousTHF (0.25 mL) and added dropwise to the nucleoside mixture. When thereaction was complete by LCMS, the reaction mixture was diluted withEtOAc and washed with saturated aqueous NaHCO₃ solution, saturated NaCl,dried over anhydrous Na₂SO₄, filtered and concentrated under reducedpressure. The residue was subjected to silica gel chromatography elutingwith 0-5% MeOH in CH₂Cl₂ followed by preparative HPLC to give theproduct (20.9 mg, 25%).

¹H NMR (300 MHz, CD₃OD) δ 7.95 (m, 1H), 7.31-6.97 (m, 7H), 4.94 (m, 1H),4.78 (m, 1H), 4.43 (m, 3H), 4.20 (m, 1H), 3.80 (d, 1H), 1.30-1.18 (m,9H);

³¹P NMR (121.4 MHz, CD₃OD) δ 3.8.

LCMS m/z 561.0 [M+H], 559.0 [M−H].

(2S)-2-ethylbutyl2-((((2R,3S,4R,5R)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-cyano-3,4-dihydroxytetrahydrofuran-2-yl)methoxy)(phenoxy)phosphorylamino)propanoate(Compound 9)

Prepared from Compound 1 and chloridate B according to the same methodas for the preparation of compound 8.

¹H NMR (300 MHz, CD₃OD) δ 7.87 (m, 1H), 7.31-7.16 (m, 5H), 6.92-6.89 (m,2H), 4.78 (m, 1H), 4.50-3.80 (m, 7H), 1.45-1.24 (m, 8H), 0.95-0.84 (m,6H).

³¹P NMR (121.4 MHz, CD₃OD) δ 3.7.

LCMS m/z 603.1 [M+H], 601.0 [M−H].

(2S)-ethyl2-((((2R,3S,4R,5R)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-cyano-3,4-dihydroxytetrahydrofuran-2-yl)methoxy)(phenoxy)phosphorylamino)propanoate(Compound 10)

Prepared from Compound 1 and chloridate A using same method as for thepreparation of compound 8.

¹H NMR (300 MHz, CD₃OD) δ 7.95 (m, 1H), 7.32-6.97 (m, 7H), 4.78 (m, 1H),4.43-4.08 (m, 6H), 3.83 (m, 1H), 1.31-1.18 (m, 6H).

³¹P NMR (121.4 MHz, CD₃OD) δ 3.7.

LCMS m/z 547.0 [M+H], 545.0 [M−H].

(2S)-ethyl2-((((2R,3R,4R,5R)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-cyano-4-fluoro-3-hydroxytetrahydrofuran-2-yl)methoxy)(phenoxy)phosphorylamino)propanoate(Compound 11)

Compound 11 was prepared from Compound 2 and chloridate A using samemethod as for the preparation of compound 8.

¹H NMR (300 MHz, CD₃OD) δ 7.91 (m, 1H), 7.33-7.16 (m, 5H), 6.98-6.90 (m,2H), 5.59 (m, 1H), 4.50-4.15 (m, 4H), 4.12-3.90 (m, 3H), 1.33-1.18 (m,6H).

³¹P NMR (121.4 MHz, CD₃OD) δ 3.8.

LCMS m/z 549.0 [M+H], 547.1 [M−H].

(2S,2′S)-diethyl2,2′-((((2R,3S,4R,5R)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-cyano-3,4-dihydroxytetrahydrofuran-2-yl)methoxy)phosphoryl)bis(azanediyl)dipropanoate(Compound 12)

The nucleoside 1 (14.6 mg, 0.05 mmol) was dissolved in anhydroustrimethyl phosphate (0.5 mL) and stirred under N₂(g) at RT. POCl₃ (9.2μL, 0.1 mmol) was added and the mixture stirred for 60 min. Alanineethyl ester hydrochloride (61 mg, 0.4 mmol) and then Et₃N (70 μL, 0.5mmol) was added. The resultant mixture was stirred for 15 min. and thenadditional Et₃N (70 μl, 0.5 mmol) was added to give a solution pH of9-10. The mixture was stirred for 2 h. and then diluted with EtOAc,washed with saturated aqueous NaHCO₃ solution followed by saturatedaqueous NaCl solution. The organic layer was dried over anhydrous Na₂SO₄and concentrated under reduced pressure. The residue was subjected topreparative HPLC (C18 column) to yield the product 12 (5.5 mg, 16%).

¹H NMR (400 MHz, CD₃OD) δ 8.13 (s, 1H), 7.41 (d, J=4.8 Hz, 1H), 7.18 (d,J=4.8 Hz, 1H), 4.78 (d, J=5.6 Hz, 1H), 4.36 (m, 1H), 4.25-4.08 (m, 7H),3.83 (m, 2H), 1.33-1.23 (m, 12H).

³¹P NMR (121.4 MHz, CD₃OD) δ 13.8.

LCMS m/z 570.0 [M+H], 568.0 [M−H].

(2S,3R,4S,5R)-2-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-2-ethynyl-5-(hydroxymethyl)tetrahydrofuran-3,4-diol(Compound 13)

The nucleoside alcohol (0.6 g, 1.08 mmol) (prepared as described inCompound 1 synthesis) was dissolved in anhydrous THF (8 mL) and placedunder N₂(g). The reaction mixture was stirred and cooled to 0° C. andthen treated with a 0.5N solution of ethynyl magnesium bromide in THF(17.2 mL, 17.2 mmol). The reaction mixture was stirred overnight at RT.AcOH (1.5 mL) was added to quench the reaction. The mixture wasconcentrated under reduced pressure and the residue redissolved inCH₂Cl₂. The solution subjected to a plug of silica gel eluting with 0 to80% EtOAc in Hexanes to provide the title product as a crude mixture.LCMS m/z 579 [M+H].

The crude ethynyl alcohol (0.624 g, 1.08 mmol) was dissolved inanhydrous CH₂Cl₂ (10 mL) and placed under N₂(g). The mixture was stirredand sulfonic acid (0.2 mL, 2.74 mmol) was added. The reaction mixturewas stirred for 12 h. at RT. When complete by LCMS, Et₃N (0.56 mL) wasadded to quench the reaction. The reaction was concentrated underreduced pressure and the residue subjected to silica gel chromatographyeluting with 0 to 75% EtOAc in Hexanes to yield the ethynyl nucleosideas a mixture of anomers (0.200 g, 33% over 2 steps). LCMS m/z 561 [M+H].

The tribenzyl nucleoside (0.650 g, 1.16 mmol) was dissolved in anhydrousCH₂Cl₂ (30 mL) and cooled to −78° C. under N₂(g). A solution of borontribromide (1 N in CH₂Cl_(2, 5.5) mL) was added and the reaction mixturestirred for 1 h. at −78° C. A solution of MeOH (10 mL) and pyridine (2mL) was added to quench the reaction and the mixture was allowed to riseto RT. The mixture was concentrated under reduced pressure and subjectedto preparative HPLC to provide the α-anomer (20 mg) and β-anomer 13 (110mg)

(β-anomer) ¹H NMR (300 MHz, DMSO) δ 7.81 (s, 1H), 7.76 (br s, 2H),6.80-6.85 (m, 2H), 5.11 (d, J=7.2 Hz, 1H), 4.90 (d, J=6.0 Hz, 1H), 4.82(dd, J=7.2, 4.8 Hz, 1H), 4.62 (t, J=6.3 Hz, 1H), 3.95-3.99 (m, 1H),3.85-3.91 (dd, J=11.4, 5.7 Hz, 1H), 3.61-3.67 (m, 1H), 3.47-3.55 (m,1H), 3.52 (d, J=0.9 Hz, 1H).

(α-anomer) ¹H NMR (300 MHz, DMSO) δ 7.80 (s, 1H), 7.59 (bs, 2H), 6.80(d, J=4.5 Hz, 1H), 6.54 (d, J=4.2 Hz, 1H), 5.00 (d, J=7.2 Hz, 1H), 4.89(d, J=4.8 Hz, 1H), 4.74 (t, J=5.7 Hz, 1H), 4.58 (t, J=4.5 Hz, 1H), 4.27(m, 1H), 3.88 (m, 1H), 3.64-3.72 (m, 1H), 3.51-3.59 (m, 1H), 3.48 (d,J=0.6 Hz, 1H)

LCMS m/z 291 [M+H].

(2R,3R,4R)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-1,3,4-tris(benzyloxy)hexane-2,5-diol(Compound 14)

The tribenzyl alcohol from Compound 1 synthesis (0.250 g, 0.453 mmol)was dissolved in anhydrous THF (25 mL) and stirred under N₂(g). Thereaction mixture was cooled to 0° C. and then a 3.0 N solution of methylmagnesium chloride in THF (1.2 mL, 3.62 mmol) was added. The reactionmixture was stirred overnight at RT. Acetic acid (1.5 mL) was added toquench the reaction and then the mixture was concentrated under reducedpressure. The residue was redissolved in CH₂Cl₂ and subjected to a plugof silica gel eluting with 0 to 80% EtOAc in hexanes. The crude product(0.452 g) was then used in the next reaction without furtherpurification.

LCMS m/z 569 [M+H].

The crude methyl nucleoside (0.452 g, 0.796 mmol) was dissolved inanhydrous CH₂Cl₂ (20 mL) and stirred under N₂(g). Methanesulfonic acid(0.2 mL, 2.78 mmol) was added and the reaction stirred for 12 hr at RT.Et₃N (0.56 mL) was added to quench the reaction and then the mixtureconcentrated under reduced pressure. The residue was subjected to silicagel chromatography eluting with 0 to 75% EtOAc in Hexanes to yield theproduct as a mixture of anomers (0.20 g, 46% over 2 steps).

LCMS m/z 551 [M+H].

The tribenzyl nucleoside (0.20 g, 0.364 mmol) was dissolved in AcOH (30mL). and charged with Pd/C (Degussa) (400 mg). The stirred mixture wasflushed with N₂(g) three times and then H₂ (g) was introduced, Thereaction was stirred under H₂ (g) for 2 h. and then the catalyst removedby filtration. The solution was concentrated under reduced pressure andunder the residue was re-dissolved in H₂O. The solution was subjected topreparative HPLC under neutral conditions to provide the α-anomer andβ-anomer 14 in 81% yield.

(α-anomer) ¹H NMR (300 MHz, D₂O) δ□□ 7.81 (s, 1H), 7.22 (d, 1H), 6.75(d, 1H), 4.47 (d, 1H), 4.25-4.31 (m, 1H), 3.88-4.95 (m, 1H), 3.58-3.86(dd, 2H), 1.50 (s, 3H).

(β-anomer) ¹H NMR (300 MHz, D₂O) δ□□ 0 7.91 (s, 1H), 7.26 (d, 1H), 6.90(d, 1H), 4.61 (d, 1H), 4.00-4.09 (m, 2H), 3.63-3.82 (dd, 2H), 1.67 (s,3H).

LCMS m/z 281 [M+H].

S,S′-2,2′-((((2R,3S,4R,5R)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-cyano-3,4-dihydroxytetrahydrofuran-2-yl)methoxy)phosphoryl)bis(oxy)bis(ethane-2,1-diyl)bis(2,2-dimethylpropanethioate) (Compound 15))

The nucleoside 1 (0.028 g, 0.096 mmol) was dissolved intrimethylphosphate (1 mL). The reaction was stirred under N₂(g) and thentreated with 1H-tetrazole (0.021 g, 0.29 mmol). The reaction mixture wascooled to 0° C. and the phosphane (Nucleoside Nucleotides, Nucleicacids; 14; 3-5; 1995; 763-766. Lefebvre, Isabelle; Pompon, Alain;Perigaud, Christian; Girardet, Jean-Luc; Gosselin, Gilles; et al.) (87mg, 0.192 mmol) was added. The reaction was stirred for 2 h. and thenquenched with 30% hydrogen peroxide (0.120 mL). The mixture was stirredfor 30 min at RT and then treated with saturated aqueous sodiumthiosulfate (1 mL). The mixture was stirred for 10 min. and thenconcentrated under reduced pressure. The residue was subjected topreparative HPLC to isolate the title product 15.

¹H NMR (300 MHz, CD₃CN) δ□□ 7.98 (s, 1H), 6.92 (d, 1H), 6.81 (d, 1H),6.44 (bs, 2H), 4.82 (m, 2H), 4.47 (m, 1H), 4.24 (m, 2H), 4.00 (m, 4H),3.80 (bs, 1H), 3.11 (m, 4H), 1.24 (s, 9H).

³¹P NMR (121.4 MHz, CD₃CN) δ −1.85 (s).

LCMS m/z 661 [M+H].

S,S′-2,2′-((((2R,3S,4R,5S)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-ethynyl-3,4-dihydroxytetrahydrofuran-2-yl)methoxy)phosphoryl)bis(oxy)bis(ethane-2,1-diyl)bis(2,2-dimethylpropanethioate) (Compound 16)

Compound 16 was prepared using the same method as compound 15 exceptsubstituting compound 13 as the starting nucleoside.

¹H NMR (300 MHz, CD₃CN) δ□□ 0 7.91 (s, 1H), 6.86 (d, J=4,8 Hz, 1H), 6.76(d, J=4.5 Hz, 1H), 6.29 (b s, 2H), 4.69 (t, J=2.7 Hz, 1H), 4.58 (d,J=5.7 Hz, 1H), 4.14-4.33 (m, 5H), 3.99-4.07 (m, 4H), 3.53 (d, J=5.4 Hz,1H), 3.11 (q, J=5.7 Hz, 4H), 1.22 (s, 18H).

LCMS m/z 658.9 [M+]. Tr=2.31

((2R,3S,4R,5R)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-cyano-3,4-dihydroxytetrahydrofuran-2-yl)methyltetrahydrogen triphosphate (Compound 17

Compound 17 was prepared from compound 1 using a similar procedure tothe preparation of compound 6. The product was isolated as the sodiumsalt.

¹H NMR (400 MHz, D₂O) δ 7.76 (s, 1H), 6.88 (d, J=4.8 Hz, 1H), 6.73 (d,J=4.4 Hz, 1H), 4.86 (d, J=5.2 Hz, 1H), 4.43 (m, 1H), 4.39 (m, 1H), 4.05(m, 1H), 3.94 (m, 1H)

³¹P NMR (121.4 MHz, D₂O) δ□ −5.4 (d, 1P), −10.8 (d, 1P), −21.1 (t, 1P).

LCMS m/z 530 [M−H], 531.9 [M+H] Tr=0.22 min

HPLC ion exchange Tr=9.95 min

((2R,3S,4R,5S)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-ethynyl-3,4-dihydroxytetrahydrofuran-2-yl)methyltetrahydrogen triphosphate (Compound 18

Compound 18 was prepared from compound 13 using a similar procedure tothe preparation of compound 6. The product was isolated as the TEA salt.

¹H NMR (300 MHz, D₂O) δ 7.85 (s, 1H), 7.09 (d, J=4.6 Hz, 1H), 6.95 (d,J=4.7 Hz, 1H), 4.23 (m, 2H), 4.08 (m, 2H), 3.06 (q, J=7.4 Hz, 20H), 1.14(t, J=7.3 Hz, 30H)

³¹P NMR (121.4 MHz, D₂O) δ□ −10.8 (d, 1P), −11.2 (d, 1P), −23.2 (t, 1P).

LCMS m/z 530.8 [M+H], Tr=0.46

HPLC ion exchange Tr=9.40 min

((2R,3S,4R,5S)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-3,4-dihydroxy-5-methyltetrahydrofuran-2-yl)methyltetrahydrogen triphosphate (Compound 19)

Compound 19 was prepared from compound 14 using a similar procedure tothe preparation of compound 6.

¹H NMR (400 MHz, D₂O) δ 7.78 (s, 1H), 6.98 (m, 1H), 6.84 (m, 1H), 4.45(m, 1H), 4.04 (m, 4H), 1.54 (s, 3H).

³¹P NMR (161 MHz, D₂O) δ −10.6 (m), −23.0 (m).

LCMS m/z 521.0 [M+H].

((2R,3R,4R,5R)-5-(4-aminopyrrolo[1,2-f][1,2,4]triazin-7-yl)-5-cyano-4-fluoro-3-hydroxytetrahydrofuran-2-yl)methyltetrahydrogen triphosphate (Compound 20)

Compound 20 was prepared from compound 2 using a similar procedure tothe preparation of compound 6.

¹H NMR (400 MHz, D₂O) δ 7.78 (s, 1H), 6.93 (d, J=4.4 Hz, 1H), 6.78 (d,J=4.8 Hz, 1H), 5.45 (dd, J=53, 4.4 Hz, 1H), 4.38-4.50 (m, 2H), 4.13-4.20(m, 2H).

³¹P NMR (161 MHz, D₂O) δ −5.7 (d, 1P), −11.0 (d, 1P), −21.5 (t, 1P).

LCMS m/z 533.9.0 [M+H], 532.0 [M−H] Tr=1.25 min.

HPLC ion exchange Tr=11.0 min

Antiviral Activity

Another aspect of the invention relates to methods of inhibiting viralinfections, comprising the step of treating a sample or subjectsuspected of needing such inhibition with a composition of theinvention.

Within the context of the invention samples suspected of containing avirus include natural or man-made materials such as living organisms;tissue or cell cultures; biological samples such as biological materialsamples (blood, serum, urine, cerebrospinal fluid, tears, sputum,saliva, tissue samples, and the like); laboratory samples; food, water,or air samples; bioproduct samples such as extracts of cells,particularly recombinant cells synthesizing a desired glycoprotein; andthe like. Typically the sample will be suspected of containing anorganism which induces a viral infection, frequently a pathogenicorganism such as a tumor virus. Samples can be contained in any mediumincluding water and organic solvent\water mixtures. Samples includeliving organisms such as humans, and man made materials such as cellcultures.

If desired, the anti-virus activity of a compound of the invention afterapplication of the composition can be observed by any method includingdirect and indirect methods of detecting such activity. Quantitative,qualitative, and semiquantitative methods of determining such activityare all contemplated. Typically one of the screening methods describedabove are applied, however, any other method such as observation of thephysiological properties of a living organism are also applicable.

The antiviral activity of a compound of the invention can be measuredusing standard screening protocols that are known. For example, theantiviral activity of a compound can be measured using the followinggeneral protocols.

Respiratory Syncytial Virus (RSV) Antiviral Activity and CytotoxicityAssays Anti-RSV Activity

Antiviral activity against RSV is determined using an in vitrocytoprotection assay in Hep2 cells. In this assay, compounds inhibitingthe virus replication exhibit cytoprotective effect against thevirus-induced cell killing that can be quantified using a cell viabilityreagent. The method used is similar to methods previously described inpublished literature (Chapman et al., Antimicrob Agents Chemother. 2007,51(9):3346-53.)

Hep2 cells are obtained from ATCC (Manassas, VI) and maintained in MEMmedia supplemented with 10% fetal bovine serum andpenicillin/streptomycin. Cells are passaged twice a week and kept atsubconfluent stage. Commercial stock of RSV strain A2 (AdvancedBiotechnologies, Columbia, Md.) is titered before compound testing todetermine the appropriate dilution of the virus stock that generatesdesirable cytopathic effect in Hep2 cells.

For antiviral tests, Hep2 cells are seeded into 96-well plates 24 hoursbefore the assay at a density of 3,000 cells/well. On a separate 96 wellplate, compounds to be tested are serially diluted in cell culturemedia. Eight concentrations in 3-fold serial dilution increments areprepared for each tested compound and 100 uL/well of each dilution istransferred in duplicate onto plates with seeded Hep2 cells.Subsequently, appropriate dilution of virus stock previously determinedby titration is prepared in cell culture media and 100 uL/well is addedto test plates containing cells and serially diluted compounds. Eachplate includes three wells of infected untreated cells and three wellsof uninfected cells that served as 0% and 100% virus inhibition control,respectively. Following the infection with RSV, testing plates areincubated for 4 days in a tissue culture incubator. After theincubation, RSV-induced cytopathic effect is determined using a CellTiterGlo reagent (Promega, Madison, Wis.) followed by a luminescenceread-out. The percentage inhibition is calculated for each testedconcentration relative to the 0% and 100% inhibition controls and theEC50 value for each compound is determined by non-linear regression as aconcentration inhibiting the RSV-induced cytopathic effect by 50%.Ribavirin (purchased from Sigma, St. Louis, Mo.) is used as a positivecontrol for antiviral activity.

Cytotoxicity

Cytotoxicity of tested compounds is determined in uninfected Hep2 cellsin parallel with the antiviral activity using the cell viability reagentin a similar fashion as described before for other cell types (Cihlar etal., Antimicrob Agents Chemother. 2008, 52(2):655-65.). The sameprotocol as for the determination of antiviral activity is used for themeasurement of compound cytotoxicity except that the cells are notinfected with RSV. Instead, fresh cell culture media (100 uL/well)without the virus is added to tested plates with cells and predilutedcompounds. Cells are then incubated for 4 days followed by a cellviability test using CellTiter Glo reagent and a luminescence read-out.Untreated cell and cells treated with 50 ug/mL puromycin (Sigma, St.Louis, Mo.) are used as 100% and 0% cell viability control,respectively. The percent of cell viability is calculated for eachtested compound concentration relative to the 0% and 100% controls andthe CC50 value is determined by non-linear regression as a compoundconcentration reducing the cell viability by 50%.

Compound EC50 / uM CC50 / uM 1 0.48 >100 10 0.18 47 12 6.5 >100 1334 >100 14 2.7 92 15 0.15 >100 16 3.3 >100

RSV RNP Preparation

RSV ribonucleoprotein (RNP) complexes were prepared from a methodmodified from Mason et al (1). HEp-2 cells were plated at a density of7.1×10⁴ cells/cm² in MEM+10% fetal bovine serum (FBS) and allowed toattach overnight at 37° C. (5% CO₂). Following attachment, the cellswere infected with RSV A2 (MOI=5) in 35 mL MEM+2% FBS. At 20 hourspost-infection, the media was replaced with MEM+2% FBS supplemented with2 μg/mL actinomycin D and returned to 37° C. for one hour. The cellswere then washed once with PBS and treated with 35 mL of PBS+250 μg/mLlyso-lecithin for one minute, after which all liquid was aspirated. Thecells were harvested by scrapping them into 1.2 mL of buffer A [50 mMTRIS acetate (pH 8.0), 100 mM potassium acetate, 1 mM DTT and 2 μg/mLactinomycin D] and lysed by repeated passage through an 18 gauge needle(10 times). The cell lysate was placed in ice for 10 minutes and thencentrifuged at 2400 g for 10 minutes at 4° C. The supernatant (S1) wasremoved and the pellet (P1) was disrupted in 600 uL of Buffer B [10 mMTRIS acetate (pH 8.0), 10 mM potassium acetate and 1.5 mM MgCl₂]supplemented with 1% Triton X-100 by repeated passage through an 18gauge needle (10 times). The resuspended pellet was placed in ice for 10minutes and then centrifuged at 2400 g for 10 minutes at 4° C. Thesupernatant (S2) was removed and the pellet (P2) was disrupted in 600 uLof Buffer B supplemented with 0.5% deoxycholate and 0.1% Tween 40. Theresuspended pellet was placed in ice for 10 minutes and then centrifugedat 2400 g for 10 minutes at 4° C. The supernatant (S3) fraction,containing the enriched RSV RNP complexes, was collected and the proteinconcentration determined by UV absorbance at 280 nm. Aliquoted RSV RNPS3 fractions were stored at −80° C.

RSV RNP Assay

Transcription reactions contained 25 μg of crude RSV RNP complexes in 30μL of reaction buffer [50 mM TRIS-acetate (pH 8.0), 120 mM potassiumacetate, 5% glycerol, 4.5 mM MgCl₂, 3 mM DTT, 2 mMethyleneglycol-bis(2-aminoethylether)-tetraacetic acid (EGTA), 50 μg/mLBSA, 2.5 U RNasin (Promega), ATP, GTP, UTP, CTP and 1.5 uCi [α-³²P] NTP(3000 Ci/mmol)]. The radiolabled nucleotide used in the transcriptionassay was selected to match the nucleotide analog being evaluated forinhibition of RSV RNP transcription. Cold, competitive NTP was added ata final concentration of one-half its K_(m) (ATP=20 μM, GTP=12.5 μM,UTP=6 μM and CTP=2 μM). The three remaining nucleotides were added at afinal concentration of 100 μM.

To determine whether nucleotide analogs inhibited RSV RNP transcription,compounds were added using a 6 step serial dilution in 5-foldincrements. Following a 90 minute incubation at 30° C., the RNPreactions were stopped with 350 μL of Qiagen RLT lysis buffer and theRNA was purified using a Qiagen RNeasy 96 kit. Purified RNA wasdenatured in RNA sample loading buffer (Sigma) at 65° C. for 10 minutesand run on a 1.2% agarose/MOPS gel containing 2M formaldehyde. Theagarose gel was dried and exposed to a Storm phosphorimager screen anddeveloped using a Storm phosphorimager (GE Healthcare). Theconcentration of compound that reduced total radiolabled transcripts by50% (IC₅₀) was calculated by non-linear regression analysis of tworeplicates.

REFERENCE

-   -   1) Mason, S., Lawetz, C., Gaudette, Y., Do, F., Scouten, E.,        Lagace, L., Simoneau, B. and Liuzzi, M. (2004)        Polyadenylation-dependent screening assay for respiratory        syncytial virus RNA transcriptase activity and identification of        an inhibitor. Nucleic Acids Research, 32, 4758-4767.

Compound IC50 / uM 6 3.6 17 1.5 18 1.6 19 1.5 20 0.8

Description of the Parainfluenza Cytoprotection Assay

The Parainfluenza Cytoprotection assay uses Vero cells and Parainfluenza3 strain C 243. Briefly virus and cells are mixed in the presence oftest compound and incubated for 7 days. The virus is pre-titered suchthat control wells exhibit 85 to 95% loss of cell viability due to virusreplication. Therefore, antiviral effect or cytoprotection is observedwhen compounds prevent virus replication. Each assay plate contains cellcontrol wells (cells only), virus control wells (cells plus virus),compound toxicity control wells (cells plus compound only), compoundcolorimetric control wells (compound only), as well as experimentalwells (compound plus cells plus virus). Cytoprotection and compoundcytotoxicity are assessed by MTS (CellTiter®96 Reagent, Promega, MadisonWis.) dye reduction. The % reduction in viral cytopathic effects (CPE)is determined and reported; IC₅₀ (concentration inhibiting virusreplication by 50%), TC₅₀ (concentration resulting in 50% cell death)and a calculated TI (therapeutic index TC₅₀/IC₅₀) are provided alongwith a graphical representation of the antiviral activity and compoundcytotoxicity when compounds are tested in dose-response. Each assayincludes ribavirin as a positive control.

Cell Preparation

Vero cells (Kidney, African green monkey, Cercopithecus aethiops) wereobtained from the American Type Culture Collection (ATCC, Rockville,Md.) and are grown in Dulbecco's Modified Eagle's Medium (DMEM)supplemented with 10% fetal bovine serum (FBS), 2.0 mM L-Glutamine, 100units/ml Penicillin and 100 ug/ml Streptomycin (“growth medium”). Cellsare sub-cultured twice a week at a split ratio of 1:10 using standardcell culture techniques. Total cell number and percent viabilitydeterminations are performed using a hemacytometer and trypan blueexclusion. Cell viability must be greater than 95% for the cells to beutilized in the assay. The cells are seeded in 96-well tissue cultureplates the day before the assay at a concentration of 1×10⁴ cells/well.

Virus Preparation

The virus used for this assay is Parainfluenza 3 strain C 243. Thisvirus was obtained from the American Type Culture Collection (ATCC) andwas grown in Vero cells for the production of stock virus pools. Foreach assay, a pre-titered aliquot of virus is removed from the freezer(−80° C.) and allowed to thaw slowly to room temperature in a biologicalsafety cabinet. The virus is resuspended and diluted into tissue culturemedium such that the amount of virus added to each well is the amountdetermined to give between 85 to 95% cell killing at 6-7 dayspost-infection.

MTS Staining for Cell Viability

At assay termination (7 days post-infection), the assay plates arestained with the soluble tetrazolium-based dye MTS(3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium;CellTiter® 96 Reagent, Promega) to determine cell viability and quantifycompound toxicity. MTS is metabolized by the mitochondrial enzymes ofmetabolically active cells to yield a soluble formazan product, allowingthe rapid quantitative analysis of cell viability and compoundcytotoxicity. This reagent is a stable, single solution that does notrequire preparation before use. At termination of the assay, 20-25 μI,of MTS reagent is added per well and the microtiter plates are thenincubated for 4-6 hrs at 37° C., 5% CO₂ to assess cell viability.Adhesive plate sealers are used in place of the lids, the sealed plateis inverted several times to mix the soluble formazan product and theplate is read spectrophotometrically at 490/650 nm with a MolecularDevices Vmax or SpectraMax Plus plate reader.

Data Analysis

Using an in-house computer program % Cytopathic Effect (CPE) Reduction,% Cell Viability, IC₂₅, IC₅₀, IC₉₅, TC₂₅, TC₅₀, and TC₉₅ and otherindices are calculated and the graphical results summary is displayed.Raw data for both antiviral activity and toxicity with a graphicalrepresentation of the data are provided in a printout summarizing theindividual compound activity. The Table below shows the activity ofselected compounds against Parainfluenza 3 virus.

Compound IC50 / uM TC50 / uM 1 1.71 >30 14 5.23 >30

The specific pharmacological and biochemical responses observed in theassays described may vary according to and depending on the particularactive compound selected or whether there are present pharmaceuticalcarriers, as well as the type of formulation and mode of administrationemployed, and such expected variations or differences in the results arecontemplated in accordance with practice of the present invention.

All publications, patents, and patent documents cited herein above areincorporated by reference herein, as though individually incorporated byreference.

The invention has been described with reference to various specific andpreferred embodiments and techniques. However, one skilled in the artwill understand that many variations and modifications may be made whileremaining within the spirit and scope of the invention.

1.-25. (canceled)
 26. A method of treating a Paramyxoviridae infectionin a mammal in need thereof, comprising administering to the mammal atherapeutically effective amount of a compound of Formula IV:

or a pharmaceutically acceptable salt, thereof; wherein: each R¹ is H orhalogen; each R³ or R⁵ is independently H, OR^(a), N(R^(a))₂, N₃, CN,NO₂, S(O)_(n)R^(a), halogen, (C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl,(C₁-C₈)substituted alkyl, (C₂-C₈)alkenyl, (C₂-C₈)substituted alkenyl,(C₂-C₈)alkynyl or (C₂-C₈)substituted alkynyl; R⁶ is OR^(a), N(R^(a))₂,N₃, CN, S(O)_(n)R^(a), —C(═O)R¹¹, —C(═O)OR¹¹, —C(═O)NR¹¹R¹², —C(═O)SR¹¹,—S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹), —S(O)₂(OR¹¹), SO₂NR¹¹R¹², halogen,(C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl, (C₁-C₈)substituted alkyl,(C₂-C₈)alkenyl, (C₂-C₈)substituted alkenyl, (C₂-C₈)alkynyl, or(C₂-C₈)substituted alkynyl; each n is independently 0, 1, or 2; eachR^(a) is independently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl,aryl(C₁-C₈)alkyl, (C₄-C₈)carbocyclylalkyl, —C(═O)R¹¹, —C(═O)OR¹¹,—C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), or —SO₂NR¹¹R¹²; R⁷ is H, —C(═O)R¹¹, —C(═O)OR¹¹,—C(═O)NR¹¹R¹², —C(═O)SR¹¹, —S(O)R¹¹, —S(O)₂R¹¹, —S(O)(OR¹¹),—S(O)₂(OR¹¹), —SO₂NR¹¹R¹², or

each Y or Y¹ is, independently, O, S, NR, ⁺N(O)(R), N(OR), ⁺N(O)(OR), orN—NR₂; W¹ and W², when taken together, are —Y³(C(R^(y))₂)₃Y³—; or one ofW¹ or W² together with either R³ or R⁴ is —Y³— and the other of W¹ or W²is Formula Ia; or W¹ and W² are each, independently, a group of theFormula Ia:

wherein: each Y² is independently a bond, O, CR₂, NR, ⁺N(O)(R), N(OR),⁺N(O)(OR), N—NR₂, S, S—S, S(O), or S(O)₂; each Y³ is independently O, S,or NR; M2 is 0, 1 or 2; each R^(x) is independently R^(y) or theformula:

wherein: each M1 a, M1 c, and M1 d is independently 0 or 1; M12c is 0,1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 or 12; each R^(y) is independently H,F, Cl, Br, I, OH, R, —C(═Y¹)R, —C(═Y¹)OR, —C(═Y¹)N(R)₂, —N(R)₂, —⁺N(R)₃,—SR, —S(O)R, —S(O)₂R, —S(O)(OR), —S(O)₂(OR), —OC(═Y¹)R, —OC(═Y¹)OR,—OC(═Y¹)(N(R)₂), —SC(═Y¹)R, —SC(═Y¹)OR, —SC(═Y¹)(N(R)₂), —N(R)C(═Y¹)R,—N(R)C(═Y¹)OR, —N(R)C(═Y¹)N(R)₂, —SO₂NR₂, —CN, —N₃, —NO₂, —OR, or W³; orwhen taken together, two R^(y) on the same carbon atom form acarbocyclic ring of 3 to 7 carbon atoms; each R is independently H,(C₁-C₈) alkyl, (C₁-C₈) substituted alkyl, (C₂-C₈)alkenyl, (C₂-C₈)substituted alkenyl, (C₂-C₈) alkynyl, (C₂-C₈) substituted alkynyl,C₆-C₂₀ aryl, C₆-C₂₀ substituted aryl, C₂-C₂₀ heterocyclyl, C₂-C₂₀substituted heterocyclyl, arylalkyl or substituted arylalkyl; W³ is W⁴or W⁵; W⁴ is R, —C(Y¹)R^(y), —C(Y¹)W⁵, —SO₂R^(y), or —SO₂W⁵; and W⁵ is acarbocycle or a heterocycle wherein W⁵ is independently substituted with0 to 3 R^(y) groups; each R⁸ is halogen, NR¹¹R¹², N(R¹¹)OR¹¹,NR¹¹NR¹¹R¹² N₃, NO, NO₂, CHO, CN, —CH(═NR¹¹), —CH═NNHR¹¹, —CH═N(OR¹¹),—CH(OR¹¹)₂, —C(═O)NR¹¹R¹², —C(═S)NR¹¹R¹², —C(═O)OR¹¹, (C₁-C₈)alkyl,(C₂-C₈)alkenyl, (C₂-C₈)alkynyl, (C₄-C₈)carbocyclylalkyl, optionallysubstituted aryl, optionally substituted heteroaryl, —C(═O)(C₁-C₈)alkyl,—S(O)_(n)(C₁-C₈)alkyl, aryl(C₁-C₈)alkyl, OR¹¹ or SR¹¹; each R⁹ isindependently H, halogen, NR¹¹R¹², N(R¹¹)OR¹¹, NR¹¹NR¹¹R¹² N₃, NO, NO₂,CHO, CN, —CH(═NR¹¹), —CH═NHNR¹¹, —CH═N(OR¹¹), —CH(OR¹¹)₂, —C(═O)NR¹¹R¹²,—C(═S)NR¹¹R¹², —C(═O)OR¹¹, R¹¹, OR¹¹ or SR¹¹; each R¹¹ or R¹² isindependently H, (C₁-C₈)alkyl, (C₂-C₈)alkenyl, (C₂-C₈)alkynyl,(C₄-C₈)carbocyclylalkyl, optionally substituted aryl, optionallysubstituted heteroaryl, —C(═O)(C₁-C₈)alkyl, —S(O)_(n)(C₁-C₈)alkyl oraryl(C₁-C₈)alkyl; or R¹¹ and R¹² taken together with a nitrogen to whichthey are both attached form a 3 to 7 membered heterocyclic ring whereinany one carbon atom of said heterocyclic ring can optionally be replacedwith —O—, —S— or —NR^(a)—; and wherein each (C₁-C₈)alkyl,(C₂-C₈)alkenyl, (C₂-C₈)alkynyl or aryl(C₁-C₈)alkyl of each R³, R⁵, R⁶,R¹¹ or R¹² is, independently, optionally substituted with one or morehalo, hydroxy, CN, N₃, N(R^(a))₂ or OR^(a); and wherein one or more ofthe non-terminal carbon atoms of each said (C₁-C₈)alkyl may beoptionally replaced with —O—, —S— or —NR^(a)—.
 27. The method of claim26 wherein each R¹, R⁵, and R⁷ is H and R³ is OR^(a).
 28. The method ofclaim 26 wherein R⁶ is CN, methyl, ethenyl, or ethynyl.
 29. The methodof claim 26 wherein R⁸ is NH₂ and R⁹ is H.
 30. The method of claim 26,wherein the compound is

pharmaceutically acceptable salt thereof.
 31. The method of claim 26,further comprising administering a pharmaceutically acceptable carrier.32. The method of claim 31 further comprising administering atherapeutically effective amount of at least one other therapeutic agentselected from the group consisting of a corticosteroid, ananti-inflammatory signal transduction modulator, β2-adrenoreceptoragonist bronchodilator, an anticholinergic, a mucolytic agent,hypertonic saline and other drugs for treating Paramyxoviridae virusinfections; or mixtures thereof.
 33. The method of claim 32 wherein theat least one other therapeutic agent is ribavirin, palivizumab,motavizumab, RSV-IGIV (RespiGam®), MEDI-557, A-60444, MDT-637 orBMS-433771 or mixtures thereof.
 34. The method of claim 26, wherein theParamyxoviridae infection is a Paramyxovirina infection.
 35. The methodof claim 34, wherein the Paramyxovirina infection is a humanparainfluenza virus infection.
 36. The method of claim 26, wherein theParamyxoviridae infection is a Pneumovirinae infection.
 37. The methodof claim 36, wherein the Pneumovirinae infection is a human respiratorysyncytial virus infection.